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WO2002011674A2 - Method and reagent for the inhibition of calcium activated chloride channel-1 (clca-1) - Google Patents

Method and reagent for the inhibition of calcium activated chloride channel-1 (clca-1) Download PDF

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WO2002011674A2
WO2002011674A2 PCT/US2001/024970 US0124970W WO0211674A2 WO 2002011674 A2 WO2002011674 A2 WO 2002011674A2 US 0124970 W US0124970 W US 0124970W WO 0211674 A2 WO0211674 A2 WO 0211674A2
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gccguuaggc cgaa
cugaugag gccguuaggc
nucleic acid
acid molecule
rna
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PCT/US2001/024970
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French (fr)
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WO2002011674A3 (en
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James Mcswiggen
James Thompson
Timothy Mckenzie
David Ayers
David E. Szymkowski
Andrew Grupe
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Ribozyme Pharmaceuticals, Incorporated
Syntex (U.S.A.) Llc
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Priority to AU2001283222A priority Critical patent/AU2001283222A1/en
Publication of WO2002011674A2 publication Critical patent/WO2002011674A2/en
Publication of WO2002011674A3 publication Critical patent/WO2002011674A3/en

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
    • C12N15/1138—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing against receptors or cell surface proteins
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/11—Antisense
    • C12N2310/111—Antisense spanning the whole gene, or a large part of it
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/12—Type of nucleic acid catalytic nucleic acids, e.g. ribozymes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/12—Type of nucleic acid catalytic nucleic acids, e.g. ribozymes
    • C12N2310/121—Hammerhead

Definitions

  • the present invention concerns compounds, compositions, and methods for the study, diagnosis, and treatment of conditions and diseases related to the expression of CLCA (Ch Channel Ca 2+ - Activated) genes.
  • CLCA proteins are emerging as a new class of channel proteins that mediate Ca 2+ -activated CI" conductance in a variety of tissues.
  • Members of the CLCA family have been cloned, isolated, and partially characterized from human, bovine, and murine species. These proteins demonstrate a high degree of homology in their size, sequence, and predicted structure yet can vary considerably in tissue distribution.
  • Bovine CLCAl (bCLCAl or CaCC) was the first reported CLCA homolog.
  • the bCLCAl protein which was isolated from and is exclusively detected in trachial epithelial cells, functions as a Ca 2+ -activated CI" channel (Ran > and Benos, 1992, J Biol.
  • Lu-ECAM-1 bovine lung-endothelial cell adhesion molecule- 1
  • Lu-ECAM-1 can mediate the binding of lung-metastatic mouse B16F10 melanoma cells to endothelial. cells (Zhu et al, 1992, J. Clin. Invest, 89, 1718-1724), however, due to sequence similarity to bCLCAl, the role of Lu-ECAM-1 as a chloride channel has been suggested (Elble et al, supra).
  • the mouse homolog, mCLCAl appears to have an expression pattern similar to the cystic fibrosis transmembrane conductance regulator (CFTR), with expression seen in various secretory epithelial cells, ' squamous epithelia, and in some lymphocytes (Gruber et al, 1998, Histochem. Cell Biol, 110, 43-49).
  • the three human CLCA homologs (hCLCAl, hCLCA2, and hCLCA3) thus far cloned, isolated, and partially characterized, all retain sequence homology, similar cDNA length, and are all located on the short arm of chromosome 1 (lp22- p31).
  • Human CLCA proteins show a restricted pattern of expression in differing secretory tissues.
  • Human CLCAl was the first reported calcium activated chloride channel in humans.
  • the 31,902-bp hCLCAl gene is located on chromosome lp22- p31, contains 14 introns, and is preceded by a canonic promoter region that contains an LI transposable element. Expression of hCLCAl is predominant in intestinal basal crypt epithelia and goblet cells.
  • hCLCAl A protein processing model has been proposed for hCLCAl in which the primary translation product (125-kDa) is cleaved to a 90- kDa and a group of 37- to 41 -kDa proteins, the latter apparently representing different glycosylation products of the same polypeptide (Gruber et al, 1998, Genomics, 54, 200-214); Transient expression of hCLCAl cDNA in HEK 293 cells is associated with an increase in whole-cell Ca 2+ -activated CI" conductance that is susceptible to inhibition with anion channel blocking compounds. Cell attached patch recordings of transfected cells in this study revealed single channels with a slope conductance of 13.4 pS (Gruber et al, supra).
  • hCLCA2 homolog is processed in a similar manner as is hCLCAl, resulting in the formation of a heterodimer consisting of a 90-kDa amino terminal and an approximately 35 -kDa carboxy terminal subunit with anchorage to the plasma membrane via four or five transmembrane domains.
  • Expression of hCLCA2 is somewhat less restricted than that of hCLCAl, being expressed from human lung, trachea, and breast tissue (Gruber et al, 1999, Am. J. Physiol, 276, C1261-C1270).
  • Human CLCA2 is expressed in normal breast epithelium but not in breast tumors of different stages of progression, suggesting that hCLCA2 may act as a tumor suppressor in breast cancer (Gruber et al, 1999, Cancer Res., 59, 5488-5491).
  • Human CLCA3 is a truncated, secreted member of the CLCA family which is expressed in numerous tissues including lung, trachea, spleen, thymus, and breast tissue.
  • hCLCA3 mRNA encodes a 37-kDa glycoprotein that corresponds to the N-terminal extracellular domain of its homologs.
  • the 37-kDa glycoprotein is secreted (Gruber and Pauli, 1999, Biochem. Biophys. Acta, 1444, 418-423).
  • the invention features novel nucleic acid-based techniques [e.g., enzymatic nucleic acid molecules (ribozymes), antisense nucleic acids, 2-5A antisense chimeras, triplex DNA, antisense nucleic acids containing RNA cleaving chemical groups] and methods for their use to modulate the expression of CLCA (CI" Channel
  • the invention features the use of one or more of the nucleic acid-based techniques independently or in combination to inhibit the expression of the genes encoding hCLCAl, hCLCA2, hCLCA3, and hCLCA4.
  • the invention features the use of nucleic acid-based techniques to specifically inhibit the expression of CLCAl (GenBank accession Nos. NM_001285, AF039400, AF039401, AF127036), CLCA2 (GenBank accession No. NM_006536), CLCA3 (GenBank accession No. NM_004921), and CLCA4 (GenBank accession No. NM_012128) genes.
  • the invention features the inhibition of CLCAl gene using the nucleic acid-based techniques of the instant invention.
  • the invention features the use of an enzymatic nucleic acid molecule, preferably in the hammerhead, NCH (hiozyme), G- cleaver, amberzyme, zinzyme and/or DNAzyme motif, to inhibit the expression of CLCA genes.
  • inhibitor it is meant that the activity of CLCAl or level of RNAs or equivalent RNAs encoding one or more protein subunits of CLCAl is reduced below that observed in the-absence of the nucleic acid molecules of the invention.
  • inhibition with enzymatic nucleic acid molecules preferably is below that level observed in the presence of an enzymatically inactive or attenuated molecule that is able to bind to the same site on the target RNA, but is unable to cleave that RNA.
  • inhibition with antisense oligonucleotides is preferably below that level observed in the presence of, for example, an oligonucleotide with scrambled sequence or with mismatches.
  • inhibition of CLCAl genes with the nucleic acid molecule of the instant invention is greater than in the presence of the nucleic acid molecule than in its absence, or the presence of a control, irrelevant, or non-inhibitory oligonucleotide.
  • enzymatic nucleic acid molecule it is meant a nucleic acid molecule which has complementarity in a substrate binding region to a specified gene target, and also has an enzymatic activity which is active to specifically cleave target RNA. That is, the enzymatic nucleic acid molecule is able to intermolecularly cleave RNA and thereby inactivate a target RNA molecule. These complementary regions allow sufficient hybridization of the enzymatic nucleic acid molecule to the target RNA and thus permit cleavage. One hundred percent complementarity is preferred, but complementarity as low as 50-75% may also be useful in this invention.
  • the nucleic acids may be modified at the base, sugar, and/or phosphate groups.
  • enzymatic nucleic acid is used interchangeably with phrases such as ribozymes, catalytic RNA, enzymatic RNA, catalytic DNA, aptazyme or aptamer-binding ribozyme, regulatable ribozyme, catalytic oligonucleotides, nucleozyme, DNAzyme, RNA enzyme, endoribonuclease, endonuclease, minizyme, leadzyme, oligozyme or DNA enzyme. All of these terminologies describe nucleic acid molecules with enzymatic activity.
  • enzymatic nucleic acid molecules described in the instant application are not meant to be limiting and those skilled in the art will recognize that all that is important in an enzymatic nucleic acid molecule of this invention is that it have a specific substrate binding site which is complementary to one or more of the target nucleic acid regions, and that it have nucleotide sequences within or surrounding that substrate binding site which impart a nucleic acid cleaving activity to the molecule (Cech et al, U.S. Patent No. 4,987,071; Cech et al, 1988, JAMA).
  • nucleic acid molecule as used herein is meant a molecule having nucleotides.
  • the nucleic acid can be single, double, or multiple stranded and may comprise modified or unmodified nucleotides or non-nucleotides or various mixtures and combinations thereof.
  • enzymatic portion or “catalytic domain” is meant that portion/region of the enzymatic nucleic acid molecule essential for cleavage of a nucleic acid substrate (for example, see Figures 1-4).
  • substrate binding arm or “substrate binding domain” is meant that portion/region of a ribozyme which is complementary to (i.e., able to base-pair with) a portion of its substrate. Generally, such complementarity is 100%, but can be less if desired. For example, as few as 10 bases out of 14 may be base-paired. Examples of such arms are shown generally in Figures 1-4. That is, these arms contain sequences within a ribozyme which are intended to bring ribozyme and target RNA together through complementary base-pairing interactions.
  • the ribozyme of the invention may have binding arms that are contiguous or non-contiguous and may be of varying lengths.
  • the length of the binding arm(s) are preferably greater than or equal to four nucleotides and of sufficient length to stably interact with the target RNA; specifically 12-100 nucleotides; more specifically 14-24 nucleotides long. If two binding arms are chosen, the design is such that the length of the binding arms are symmetrical (i.e., each of the binding arms is of the same length; e.g., five and five nucleotides, six and six nucleotides or seven and seven nucleotides long) or asymmetrical (i.e., the binding arms are of different length; e.g., six and three nucleotides; three and six nucleotides long; four and five nucleotides long; four and six nucleotides long; four and seven nucleotides long; and the like).
  • NCH or "Inozyme” motif is meant, an enzymatic nucleic acid molecule comprising a motif as described in Ludwig et al, USSN No. 09/406,643, filed September 27, 1999, entitled “COMPOSITIONS HAVING RNA CLEANING ACTIVITY", and International PCT publication Nos. WO 98/58058 and WO 98/58057, all incorporated by reference herein in their entirety including the drawings.
  • G-cleaver an enzymatic nucleic acid molecule comprising a motif as described in Eckstein et al, mtemational PCT publication No. WO 99/16871, incorporated by reference herein in its entirety including the drawings.
  • zinzyme motif is meant, a class ⁇ enzymatic nucleic acid molecule comprising a motif as described in Beigelman et al, International PCT publication
  • Zinzymes represent a non-limiting example of an enzymatic nucleic acid molecule that does not require a ribonucleotide (2' -OH) group within its own nucleic
  • Amberzyme motif is meant, a class I enzymatic nucleic acid molecule comprising a motif as described in Beigelman et al, International PCT publication No. WO 99/55857, incorporated by reference herein in its entirety including the drawings.
  • Amberzymes represent a non-limiting example of an enzymatic nucleic acid molecule that does not require a ribonucleotide (2'-OH) group within its own nucleic acid sequence for activity.
  • DNAzyme' is meant, an enzymatic nucleic acid molecule that does not require the presence of a ribonucleotide (2' -OH) group within the DNAzyme molecule for its activity.
  • the enzymatic nucleic acid molecule may have an attached linker(s) or other attached or associated groups, moieties, or chains containing one or more nucleotides with 2'-OH groups.
  • DNAzyme can be synthesized chemically or expressed endogenously in vivo, by means of a single stranded DNA vector or equivalent thereof.
  • sufficient length is meant an oligonucleotide of greater than or equal to 3 nucleotides that is of a length great enough to provide the intended function under the expected condition.
  • sufficient length means that the binding arm sequence is long enough to provide stable binding to a target site under the expected binding conditions. Preferably, the binding arms are not so long as to prevent useful turnover.
  • stably interact is meant, interaction of the oligonucleotides with target nucleic acid (e.g., by forming hydrogen bonds with complementary nucleotides in the target under physiological conditions).
  • RNA to CLCAl is meant to include those naturally occurring RNA molecules having homology (partial or complete) to CLCAl proteins or encoding for proteins with similar function as CLCAl in various organisms, including human, rodent, primate, rabbit, pig, protozoans, fungi, plants, and other microorganisms and parasites.
  • the equivalent RNA sequence also includes in addition to the coding region, regions such as 5 '-untranslated region, 3 '-untranslated region, introns, intron-exon junction and the like.
  • nucleotide sequence of two or more nucleic acid molecules is partially or completely identical.
  • antisense nucleic acid a non-enzymatic nucleic acid molecule that binds to target RNA by means of RNA-RNA or RNA-DNA or RNA- PNA (protein nucleic acid; Egholm et al, 1993 Nature 365, 566) interactions and alters the activity of the target RNA (for a review, see Stein and Cheng, 1993 Science 261, 1004 and Woolf et al, US patent No. 5,849,902).
  • antisense molecules will be complementary to a target sequence along a single contiguous sequence of the antisense molecule.
  • an antisense molecule may bind to substrate such that the substrate molecule forms a loop, and/or an antisense molecule may bind such that the antisense molecule forms a loop.
  • the antisense molecule may be complementary to two (or even more) noncontiguous substrate sequences or two (or even more) non-contiguous sequence portions of an antisense molecule may be complementary to a target sequence or both.
  • antisense DNA can be used to target RNA by means of DNA-RNA interactions, thereby activating RNase H, which digests the target RNA in the duplex.
  • Antisense DNA can be synthesized chemically or expressed via the use of a single stranded DNA expression vector or equivalent thereof.
  • 2-5A antisense chimera an antisense oligonucleotide containing a 5'-phosphorylated 2'-5'-linked adenylate residue. These chimeras bind to target RNA in a sequence-specific manner and activate a cellular 2-5A-dependent ribonuclease which, in turn, cleaves the target RNA (Torrence et al, 1993 Proc.
  • triplex DNA it is meant an oligonucleotide that can bind to a double- stranded DNA in a sequence-specific manner to form a triple-strand helix. Formation of such triple helix structure has been shown to inhibit transcription of the targeted gene (Duval- Valentin et al, 1992 Proc. Natl Acad. Sci. USA 89, 504).
  • RNA By “gene” it is meant a nucleic acid that encodes an RNA.
  • nucleic acid can form hydrogen bond(s) with another RNA sequence by either traditional Watson-Crick or other non-traditional types.
  • the binding free energy for a nucleic acid molecule with its target or complementary sequence is sufficient to allow the relevant function of the nucleic acid to proceed, e.g., ribozyme cleavage, antisense or triple helix inhibition. Determination of binding free energies for nucleic acid molecules is well known in the art (see, e.g., Turner et al., 1987, CSHSymp. Quant. Biol. UL pp.123-133; Frier et al, 1986, Proc. Nat. Acad.
  • a percent complementarity indicates the percentage of contiguous residues in a nucleic acid molecule which can form hydrogen bonds (e.g., Watson- Crick base pairing) with a second nucleic acid sequence (e.g., 5, 6, 7, 8, 9, 10 out of 10 being 50%, 60%, 70%, 80%, 90%, and 100% complementary).
  • Perfectly complementary means that all the contiguous residues of a nucleic acid sequence will hydrogen bond with the same number of contiguous residues in a second nucleic acid sequence.
  • enzymatic nucleic acids act by first binding to a target RNA. Such binding occurs through the target binding portion of a enzymatic nucleic acid which is held in close proximity to an enzymatic portion of the molecule that acts to cleave the target RNA.
  • the enzymatic nucleic acid first recognizes and then binds a target RNA through complementary base-pairing, and once bound to the correct site, acts enzymatically to cut the target RNA. Strategic cleavage of such a target RNA will destroy its ability to direct synthesis of an encoded protein. After an enzymatic nucleic acid has bound and cleaved its RNA target, it is released from that RNA to search for another target and can repeatedly bind and cleave new targets. Thus, a single ribozyme molecule is able to cleave many molecules of target RNA.
  • the ribozyme is a highly specific inhibitor of gene expression, with the specificity of inhibition depending not only on the base-pairing mechanism of binding to the target RNA, but also on the mechanism of target RNA cleavage. Single mismatches, or base-substitutions, near the site of cleavage can completely eliminate catalytic activity of a ribozyme.
  • the enzymatic nucleic acid molecule that cleave the specified sites in CLCAl - specific RNAs represent a novel therapeutic approach to treat Chronic Obstructive
  • COPDs Pulmonary Diseases
  • chronic bronchitis chronic bronchitis
  • asthma chronic bronchitis
  • cystic fibrosis chronic fibrosis
  • obstructive bowel syndrome and other indications that may respond to the level of
  • the enzymatic nucleic acid molecule is formed in a hammerhead or hairpin motif, but may also be formed in the motif of a hepatitis delta vims, group I intron, group ⁇ intron or RNase P RNA (in association with an RNA guide sequence), Neurospora VS RNA, DNAzymes, NCH cleaving motifs, or G-cleavers.
  • hammerhead motifs are described by Dreyfus, supra, Rossi et al, 1992, AIDS Research and Human Retroviruses 8, 183;
  • hairpin motifs are described by Hampel et ah, EP0360257, Hampel and Tritz, 1989 Biochemistry 28, 4929, Feldstein et al, 1989, Gene 82, 53, Haseloff and Gerlach, 1989, Gene, 82, 43, Hampel et al, 1990 Nucleic Acids Res. 18, 299; Chowrira & McSwiggen, US. Patent No. 5,631,359.
  • the hepatitis delta vims motif is described by Perrotta and Been, 1992 Biochemistry 31, 16.
  • the RNase P motif is described by Guerrier- Takada et al, 1983 Cell 35, 849; Forster and Altman, 1990, Science 249, 783; Li and Altman, 1996, Nucleic Acids Res. 24, 835.
  • Neurospora VS RNA ribozyme motif is described by Collins (Saville and Collins, 1990 Cell 61, 685-696; Saville and Collins, 1991 Proc. Natl. Acad. Sci. USA 88, 8826-8830; Collins and Olive, 1993 Biochemistry 32, 2795-2799; Guo and Collins, 1995, EMBO. J. 14, 363).
  • Group II introns are described by Griffin et al, 1995, Chem. Biol.
  • WO 98/58058 and G-cleavers are described in Kore et al., 1998, Nucleic Acids Research 26, 4116-4120 and Eckstein et al, International PCT Publication No. WO 99/16871. Additional motifs such as the Aptazyme (Breaker et al, WO 98/43993), Amberzyme (Class I motif; Figure 3; Beigelman et al, International PCT publication No. WO 99/55857) and Zinzyme (Beigelman et al, International PCT publication No. WO 99/55857), all these references are incorporated by reference herein in their totalities, including drawings and can also be used in the present invention.
  • a nucleic acid molecule e.g., an antisense molecule, a triplex DNA, or a ribozyme
  • a nucleic acid molecule is 13 to 100 nucleotides in length, e.g., in specific embodiments 35, 36, 37, or 38 nucleotides in length (e.g., for particular ribozymes or. antisense).
  • the nucleic acid molecule is 15-100, 17-100, 20-100, 21-100, 23-100, 25-100, 27-100, 30-100, 32- 100, 35-100, 40-100, 50-100, 60-100, 70-100, or 80-100 nucleotides in length.
  • the upper limit of the length range can be, for example, 30, 40, 50, 60, 70, or 80 nucleotides.
  • the length range for particular embodiments has lower limit as specified, with an upper limit as specified which is greater than the lower limit.
  • the length range can be 35-50 nucleotides in length. All such ranges are expressly included.
  • a nucleic acid molecule can have a length which is any of the lengths specified above, for example, 21 nucleotides in length.
  • the invention provides a method for producing a class of nucleic acid-based gene inhibiting agents which exhibit a high degree of specificity for the RNA of a desired target.
  • the enzymatic nucleic acid molecule is preferably targeted to a highly conserved sequence region of target RNAs encoding CLCA proteins (for example, CLCAl, CLCA2, CLCA3 and/or CLCA4) such that specific treatment of a disease or condition can be provided with either one or several nucleic acid molecules of the invention.
  • Such nucleic acid molecules can be delivered exogenously to specific tissue or cellular targets as required.
  • the nucleic acid molecules e.g., ribozymes and antisense
  • the invention features the use of nucleic acid-based inhibitors of the invention to specifically target genes that share homology with the CLCAl gene.
  • cell is used in its usual biological sense, and does not refer to an entire multicellular organism, e.g., specifically does not refer to a human.
  • the cell may be present in a non-human multicellular organism, e.g., birds, plants and mammals such as cows, sheep, apes, monkeys, swine, dogs, and cats.
  • CLCA proteins is meant, a protein or a mutant protein derivative thereof, comprising a calcium activated chloride channel protein.
  • highly conserved sequence region a nucleotide sequence of one or more regions in a target gene does not vary significantly from one generation to the other or from one biological system to the other.
  • the nucleic acid-based inhibitors of CLCAl expression are useful for the prevention and/or treatment of diseases and conditions including Chronic Obstructive Pulmonary Disease (COPD), chronic bronchitis, asthma, cystic fibrosis, obstructive bowel syndrome, and any other diseases or conditions that are related to or will respond to the levels of CLCAl in a cell or tissue, alone or in combination with other therapies.
  • COPD Chronic Obstructive Pulmonary Disease
  • COPD Chronic Obstructive Pulmonary Disease
  • chronic bronchitis asthma
  • cystic fibrosis obstructive bowel syndrome
  • any other diseases or conditions that are related to or will respond to the levels of CLCAl in a cell or tissue, alone or in combination with other therapies.
  • the nucleic acid-based inhibitors of the invention are added directly, or can be complexed with cationic lipids, packaged within liposomes, or otherwise delivered to target cells or tissues.
  • the nucleic acid or nucleic acid complexes can be locally administered to relevant tissues ex vivo, or in vivo through injection, infusion pump or stent, with or without their incorporation in biopolymers.
  • the enzymatic nucleic acid inhibitors comprise sequences, which are complementary to the substrate sequences in Tables III to IX. Examples of such enzymatic nucleic acid molecules also are shown in Tables III to IX. Examples of such enzymatic nucleic acid molecules consist essentially of sequences defined in these Tables.
  • the invention features antisense nucleic acid molecules and 2-5A chimera including sequences complementary to the substrate sequences shown in Tables III to IX.
  • nucleic acid molecules can include sequences as shown for the binding arms of the enzymatic nucleic acid molecules in Tables III to VIII and sequences shown as GeneBlocTM sequences in Table IX.
  • triplex molecules can be provided targeted to the corresponding DNA target regions, and containing the DNA equivalent of a target sequence or a sequence complementary to the specified target (substrate) sequence.
  • antisense molecules will be complementary to a target sequence along a single contiguous sequence of the antisense molecule.
  • an antisense molecule may bind to substrate such that the substrate molecule forms a loop, and/or an antisense molecule may bind such that the antisense molecule forms a loop.
  • the antisense molecule may be complementary to two (or even more) non- contiguous substrate sequences or two (or even more) non-contiguous sequence portions of an antisense molecule may be complementary to a target sequence or both.
  • the active nucleic acid molecule of the invention for example, an enzymatic nucleic acid molecule, contains an enzymatic center or core equivalent to those in the examples, and binding arms able to bind RNA such that cleavage at the target site occurs.
  • a core region can, for example, include one or more loop, stem-loop structure, or linker which does not prevent enzymatic activity.
  • the underlined regions in the sequences in Tables III, IV and VIII can be such a loop, stem-loop, nucleotide linker, and/or non- nucleotide linker and can be represented generally as sequence "X".
  • a core sequence for a hammerhead enzymatic nucleic acid can comprise a conserved sequence, such as 5'-CUGAUGAG-3' and 5'-CGAA-3' connected by "X", where X is 5'-GCCGUUAGGC-3' (SEQ ID NO 5450), or any other Stem ⁇ region known in the art.
  • ribozymes or antisense molecules that interact with target RNA molecules and inhibit CLCAl (specifically CLCAl gene) activity are expressed from transcription units inserted into DNA or RNA vectors.
  • the recombinant vectors are preferably DNA plasmids or viral vectors.
  • Ribozyme or antisense expressing viral vectors could be constracted based on, but not limited to, adeno-associated virus, retrovirus, adenovirus, or alphavirus.
  • the recombinant vectors capable of expressing the ribozymes or antisense are delivered as described above, and persist in target cells.
  • viral vectors may be used that provide for transient expression of ribozymes or antisense. Such vectors can be repeatedly administered as necessary. Once expressed, the ribozymes or antisense bind to the target RNA and inhibit its function or expression.
  • ribozyme or antisense expressing vectors can be systemic, such as by intravenous or intramuscular administration, by administration to target cells ex-planted from the patient followed by reintroduction into the patient, or by any other means that would allow for introduction into the desired target cell.
  • Antisense DNA can be expressed endogenously via the use of a single stranded DNA intracellular expression vector.
  • RNA is meant a molecule comprising at least one ribonucleotide residue.
  • ribonucleotide is meant a nucleotide with a hydroxyl group at the 2' position of a ⁇ -D-ribo-furanose moiety.
  • vectors any nucleic acid- and/or viral-based technique used to deliver a desired nucleic acid.
  • patient is meant an organism, which is a donor or recipient of explanted cells or the cells themselves.
  • patient also refers to an organism to which the nucleic acid molecules of the invention can be administered.
  • a patient is a mammal or mammalian cells. More preferably, a patient is a human or human cells.
  • nucleic acid molecules of the instant invention can be used to treat diseases or conditions discussed above.
  • the patient may be treated, or other appropriate cells may be treated, as is evident to those skilled in the art, individually or in combination with one or. more drugs under conditions suitable for the treatment.
  • the described molecules can be used in combination with other known treatments to treat conditions or diseases discussed above.
  • the described molecules could be used in combination with one or more known therapeutic agents to treat Chronic Obstructive Pulmonary Diseases (COPDs), chronic bronchitis, asthma, cystic fibrosis, obstructive bowel syndrome, and/or other disease states or conditions which respond to the modulation of CLCAl expression.
  • COPDs Chronic Obstructive Pulmonary Diseases
  • chronic bronchitis chronic bronchitis
  • asthma cystic fibrosis
  • obstructive bowel syndrome obstructive bowel syndrome
  • the invention features nucleic acid-based inhibitors (e.g., enzymatic nucleic acid molecules (ribozymes), antisense nucleic acids, 2-5A antisense chimeras, triplex DNA, antisense nucleic acids containing RNA cleaving chemical groups) and methods for their use to down regulate or inhibit the expression of genes (e.g., CLCAl) capable of progression and/or maintenance of Chronic Obstructive Pulmonary Diseases (COPDs), chronic bronchitis, asthma, cystic fibrosis, obstructive bowel syndrome, and/or other disease states or conditions which respond to the modulation of CLCAl expression.
  • genes e.g., CLCAl
  • COPDs Chronic Obstructive Pulmonary Diseases
  • COPDs Chronic Obstructive Pulmonary Diseases
  • Figure 1 shows examples of chemically stabilized ribozyme motifs.
  • HH Rz represents hammerhead ribozyme motif (Usman et al, 1996, Curr. Op. Struct. Bio., 1, 527);
  • NCH Rz represents the NCH ribozyme motif (Ludwig & Sproat, Intemational PCT Publication No. WO 98/58058);
  • G-CIeaver represents G-cleaver ribozyme motif (Kore et al, 1998, Nucleic Acids Research 26, 4116-4120).
  • N or n represent independently a nucleotide which may be same or different and have complementarity to each other; ri, represents ribo-Inosine nucleotide; arrow indicates the site of cleavage within the target.
  • Position 4 of the HH Rz and the NCH Rz is shown as having 2'-C-allyl modification, but those skilled in the art will recognize that this position can be modified with other modifications well known in the art, so long as such modifications do not significantly inhibit the activity of the ribozyme.
  • Figure 2 shows an example of the Amberzyme ribozyme motif that is chemically stabilized (see, for example, Beigelman et al, Intemational PCT publication No. WO 99/55857, incorporated by reference herein; also referred to as
  • the Amberzyme motif is a class of enzymatic nucleic molecules that do not require the presence of a ribonucleotide (2'-OH) group for its activity.
  • Figure 3 shows an example of the Zinzyme A ribozyme motif that is chemically stabilized (Beigelman et al, International PCT publication No. WO
  • the Zinzyme motif is a class of enzymatic nucleic molecules that do not require the presence of a ribonucleotide (2' -OH) group for its activity.
  • Figure 4 shows an example of a DNAzyme motif described by Santoro et al., 1991, PNAS, 94, 4262.
  • Figures 5A and 5B are diagrammatic schemes representative of the process used for Target Discovery in the instant invention.
  • the process for Target Discovery is described in Jarvis et al, Intemational PCT publication No. WO 98/50530, incorporated by reference herein in its entirety including the Figures.
  • Antisense molecules may be modified or unmodified RNA, DNA, or mixed polymer oligonucleotides which primarily function by specifically binding to matching sequences resulting in inhibition of peptide synthesis (Wu-Pong, Nov 1994, BioPharm, 20-33).
  • the antisense oligonucleotide binds to target RNA by Watson Crick base-pairing and blocks gene expression by preventing ribosomal translation of the bound sequences either by steric blocking or by activating RNase H enzyme.
  • Antisense molecules can also alter protein synthesis by interfering with RNA processing or transport from the nucleus into the cytoplasm (Mukhopadhyay & Roth, 1996, Crit. Rev. in Oncogenesis 7, 151-190).
  • binding of single stranded DNA to RNA may result in nuclease degradation of the heteroduplex (Wu-Pong, supra; Crooke, supra).
  • the only backbone modified DNA chemistry which will act as substrates for RNase H are phosphorothioates, phosphorodithioates, and borontrifluoridates.
  • 2'-arabino and 2'-fluoro arabino- containing oligos can also activate RNase H activity.
  • antisense molecules have been described that utilize novel configurations of chemically modified nucleotides, secondary stracture, and/or RNase H substrate domains (Woolf et al, International PCT Publication No. WO 98/13526; Thompson et al, Intemational PCT Publication No. WO 99/54459; Hartmann et al, USSN 60/101,174 which was filed on September 21, 1998) all of these are incorporated by reference herein in their entirety.
  • antisense deoxyoligoribonucleotides can be used to target RNA by means of DNA-RNA interactions, thereby activating RNase H, which digests the target RNA in the duplex.
  • Antisense DNA can be expressed endogenously in vivo via the use of a single stranded DNA intracellular expression vector or equivalents and variations thereof.
  • TFO Triplex Forming Oligonucleotides
  • Single stranded DNA may be designed to bind to genomic DNA in a sequence specific manner.
  • TFOs are comprised of pyrimidine-rich oligonucleotides which bind DNA helices through Hoogsteen Base-pairing (Wu-Pong, supra). The resulting triple helix composed of the DNA sense, DNA antisense, and TFO disrupts RNA synthesis by RNA polymerase.
  • the TFO mechanism may result in gene expression or cell death since binding may be irreversible (Mukhopadhyay & Roth, supra).
  • 2-5A Antisense Chimera The 2-5A system is an interferon mediated mechanism for RNA degradation found in higher vertebrates (Mitra et al, 1996, Proc Nat Acad Sci USA 93, 6780-6785). Two types of enzymes, 2-5A synthetase and RNase L, are required for RNA cleavage.
  • the 2-5A synthetases require double stranded RNA to form 2'-5' oligoadenylates (2-5 A).
  • 2-5 A then acts as an allosteric effector for utilizing RNase L which has the ability to cleave single stranded RNA.
  • the ability to form 2-5A structures with double stranded RNA makes this system particularly useful for inhibition of viral replication.
  • (2'-5') oligoadenylate structures may be covalently linked to antisense molecules to form chimeric oligonucleotides capable of RNA cleavage (Torrence, supra). These molecules putatively bind and activate a 2-5 A dependent RNase, the oligonucleotide/enzyme complex then binds to a target RNA molecule which can then be cleaved by the RNase enzyme.
  • Enzymatic Nucleic Acid Seven basic varieties of naturally occurring enzymatic RNAs are presently known.
  • several in vitro selection (evolution) strategies have been used to evolve new nucleic acid catalysts capable of catalyzing cleavage and ligation of phosphodiester linkages (Joyce, 1989, Gene, 82, 83-87; Beaudry et al, 1992, Science 257, 635-641; Joyce, 1992, Scientific American 267, 90-97; Breaker et al, 1994, TIBTECH 12, 268; Bartel et al., 1993, Science 261:1411-1418; Szostak, 1993, IWS 17, 89-93; Kumar et al, 1995, FASEB J., 9, 1183; Breaker, 1996, Curr.
  • Nucleic acid molecules of this invention will block to some extent CLCAl protein expression and can be used to treat disease or diagnose disease associated with the levels of CLCAl.
  • ribozyme has significant advantages, such as the concentration of ribozyme necessary to affect a therapeutic treatment is lower. This advantage reflects the ability of the ribozyme to act enzymatically.
  • a single ribozyme molecule is able to cleave many molecules of target RNA.
  • the ribozyme is a highly specific inhibitor, with the specificity of inhibition depending not only on the base-pairing mechanism of binding to the target RNA, but also on the mechanism of target RNA cleavage. Single mismatches, or base-substitutions, near the site of cleavage can be chosen to completely eliminate catalytic activity of a ribozyme.
  • Nucleic acid molecules having an endonuclease enzymatic activity are able to repeatedly cleave other separate RNA molecules in a nucleotide base sequence- specific manner. Such enzymatic nucleic acid molecules can be targeted to virtually any RNA transcript, and achieve efficient cleavage in vitro (Zaug et al, 324, Nature 429 1986 ; Uhlenbeck, 1987 Nature 328, 596; Kim et al, 84 Proc. Natl Acad. Sci. USA 8788, 1987; Dreyfus, 1988, Einstein Quart. J. Bio.
  • Ribozymes can be designed to cleave specific RNA targets within the background of cellular RNA. Such a cleavage event renders the RNA nonfunctional and abrogates protein expression from that RNA. In this manner, synthesis of a protein associated with a disease state can be selectively inhibited (Warashina et al, 1999, Chemistry and Biology, 6, 237-250).
  • the nucleic acid molecules of the instant invention are also referred to as GeneBloc reagents, which are essentially nucleic acid molecules (e.g.; ribozymes, antisense) capable of down-regulating gene expression.
  • GeneBlocs are modified oligonucleotides including ribozymes and modified antisense oligonucleotides that bind— to and target specific mRNA molecules. Because GeneBlocs can be designed to target any specific mRNA, their potential applications are quite broad. Traditional antisense approaches have often relied heavily on the use of phosphorothioate modifications to enhance stability in biological samples, leading to a myriad of specificity problems stemming from nonspecific protein binding and general cytotoxicity (Stein, 1995, Nature Medicine, 1, 1119).
  • GeneBlocs contain a number of modifications that confer nuclease resistance while making minimal use of phosphorothioate linkages, which reduces toxicity, increases binding affinity and minimizes non-specific effects compared with traditional antisense oligonucleotides. Similar reagents have recently been utilized successfully in various cell culture systems (Vassar, et al, 1999, Science, 286, 735) and in vivo (Jarvis et al., manuscript in preparation). In addition, novel cationic lipids can be utilized to enhance cellular uptake in the presence of serum.
  • Targets for useful ribozymes and antisense nucleic acids can be determined as disclosed in Draper et al, WO 93/23569; Sullivan et al, WO 93/23057; Thompson et al, WO 94/02595; Draper et al, WO 95/04818; McSwiggen et al, US Patent No. 5,525,468. All of these publications are hereby incorporated by reference herein in their totality. Other examples include the following PCT applications, which concern inactivation of expression of disease-related genes: WO 95/23225, WO 95/13380, WO 94/02595, all of which are incorporated by reference herein.
  • Ribozymes and antisense to such targets are designed as described in those applications and synthesized to be tested in vitro and in vivo, as also described.
  • the sequences of human CLCAl RNAs were screened for optimal enzymatic nucleic acid and antisense target sites using a computer-folding algorithm. Antisense, hammerhead, DNAzyme, NCH, amberzyme, zinzyme, or G-Cleaver ribozyme binding/cleavage sites were identified.
  • Antisense, hammerhead, DNAzyme, NCH, amberzyme, zinzyme or G-Cleaver ribozyme binding/cleavage sites were identified.
  • the nucleic acid molecules are individually analyzed by computer folding (Jaeger et al, 1989 Proc. Natl Acad. Sci. USA, 86, 7706) to assess whether the sequences fold into the appropriate secondary stracture. Those nucleic acid molecules with unfavorable intramolecular interactions such as between the binding arms and the catalytic core are eliminated from consideration. Varying binding arm lengths can be chosen to optimize activity.
  • Antisense, hammerhead, DNAzyme, NCH, amberzyme, zinzyme or G-Cleaver ribozyme binding/cleavage sites were identified and were designed to anneal to various sites in the RNA target.
  • the binding arms are complementary to the target site sequences described above.
  • the nucleic acid molecules were chemically synthesized. The method of synthesis used follows the procedure for normal DNA/RNA synthesis as described below and in Usman et al, 1987 J. Am. Chem. Soc, 109, 7845; Scaringe et al, 1990 Nucleic Acids Res., 18, 5433; Wincott et al, 1995 Nucleic Acids Res. 23, 2677-2684; and Carafhers et al., 1992, Methods in Enzymology 211,3-19.
  • nucleic acids greater than 100 nucleotides in length is difficult using automated methods, and the therapeutic cost of such molecules is prohibitive.
  • small nucleic acid motifs (“small refers to nucleic acid motifs no more than 100 nucleotides in length, preferably no more than 80 nucleotides in length, and most preferably no more than 50 nucleotides in length; e.g., antisense oligonucleotides, hammerhead or the NCH ribozymes) are preferably used for exogenous delivery.
  • the simple structure of these molecules increases the ability of the nucleic acid to invade targeted regions of RNA structure.
  • Exemplary molecules of the instant invention are chemically synthesized, and others can similarly be synthesized.
  • Oligonucleotides are synthesized using protocols known in the art as described in Carathers et al, 1992, Methods in Enzymology 211, 3-19, Thompson et al, Intemational PCT Publication No. WO 99/54459, Wincott et al, 1995, Nucleic Acids Res. 23, 2677-2684, Wincott et al, 1997, Methods Mol. Bio., 74, 59, Brennan et al, 1998, Biotechnol Bioeng., 61, 33-45, and Brennan, US patent No. 6,001,311. All of these references are incorporated herein by reference.
  • oligonucleotides makes use of common nucleic acid protecting and coupling groups, such as dimethoxytrityl at the 5'-end, and phosphoramidites at the 3'-end.
  • small scale syntheses are conducted on a 394 Applied Biosystems, Inc. synthesizer using a 0.2 ⁇ mol scale protocol with a 2.5 min coupling step for 2'-O-methylated nucleotides and a 45 sec coupling step for 2'- deoxy nucleotides.
  • Table II outlines the amounts and the contact times of the reagents used in the synthesis cycle.
  • syntheses at the 0.2 ⁇ mol scale can be performed on a 96-well plate synthesizer, such as the instrument produced by Protogene (Palo Alto, CA) with minimal modification to the cycle.
  • synthesizer include; detritylation solution is 3% TCA in methylene chloride (ABI); capping is performed with 16% N-methyl imidazole in THF (ABI) and 10% acetic anhydride/10% 2,6-lutidine in THF (ABI); and oxidation solution is 16.9 mM 12, 49 mM pyridine, 9% water in THF (PERSEPTIVETM). Burdick & Jackson Synthesis Grade acetonitrile is used directly from the reagent bottle. S-Ethyltetrazole solution (0.25 M in acetonitrile) is made up from the solid obtained from American International Chemical, Inc.
  • Beaucage reagent (3H-l,2-Benzodithiol-3-one 1,1 -dioxide, 0.05 M in acetonitrile) is used.
  • Deprotection of the antisense oligonucleotides is performed as follows: the polymer-bound trityl-on oligoribonucleotide is transferred to a 4 mL glass screw top vial and suspended in a solution of 40% aq. methylamine (1 mL) at 65 °C for 10 min. After cooling to -20 °C, the supernatant is removed from the polymer support. The support is washed three times with 1.0 mL of EtOH:MeC ⁇ :H2O/3:l:l, vortexed and the supernatant is then added to the first supernatant. The combined supernatants, containing the oligoribonucleotide, are dried to a white powder.
  • RNA including certain enzymatic nucleic acid molecules follows the procedure as described in Usman et al, 1987, J. Am. Chem. Soc, 109, 7845; Scaringe et al, 1990, Nucleic Acids Res., 18, 5433; Wincott et al, 1995, Nucleic Acids Res. 23, 2677-2684 and Wincott et al, 1997, Methods Mol. Bio., 74, 59, and makes use of common nucleic acid protecting and coupling groups, such as dimethoxytrityl at the 5'-end, and phosphoramidites at the 3'-end.
  • common nucleic acid protecting and coupling groups such as dimethoxytrityl at the 5'-end, and phosphoramidites at the 3'-end.
  • small scale syntheses are conducted on a 394 Applied Biosystems, Inc. synthesizer using a 0.2 ⁇ mol scale protocol with a 7.5 min coupling step for alkylsilyl protected nucleotides and a 2.5 min coupling step for 2'- O-methylated nucleotides.
  • Table II outlines the amounts and the contact times of the reagents used in the synthesis cycle.
  • syntheses at the 0.2 ⁇ mol scale can be done on a 96-well plate synthesizer, such as the instrument produced by Protogene (Palo Alto, CA) with minimal modification to the cycle.
  • Average coupling yields on the 394 Applied Biosystems, Inc. synthesizer, determined by colorimetric quantitation of the trityl fractions, are typically 97.5- 99%.
  • synthesizer include; detritylation solution is 3% TCA in methylene chloride (ABI); capping is performed with 16% N-methyl imidazole in THF (ABI) and 10% acetic anhydride/10% 2,6-lutidine in THF (ABI); oxidation solution is 16.9 mM ⁇ 2, 49 mM pyridine, 9% water in THF (PERSEPTIVETM). Burdick & Jackson Synthesis Grade acetonitrile is used directly from the reagent bottle. S-Ethyltetrazole solution (0.25 M in acetonitrile) is made up from the solid obtained from American Intemational Chemical, Inc.
  • Beaucage reagent (3H-l,2-Benzodifhiol-3-one l,l-dioxide0.05 M in acetonitrile) is used.
  • Deprotection of the R ⁇ A is performed using either a two-pot or one-pot protocol.
  • the polymer-bound trityl-on oligoribonucleotide is transferred to a 4 mL glass screw top vial and suspended in a solution of 40% aq. methylamine (1 mL) at 65 °C for 10 min. After cooling to -20 °C, the supernatant is removed from the polymer support. The support is washed three times with 1.0 mL of EtOH:MeC ⁇ :H2O/3:l:l, vortexed and the supernatant is then added to the first supernatant.
  • the combined supernatants, containing the oligoribonucleotide, are dried to a white powder.
  • the base deprotected oligoribonucleotide is resuspended in anhydrous TEA HF/NMP solution (300 ⁇ L of a solution of 1.5 mL N- methylpyrrolidinone, 750 ⁇ L TEA and 1 mL TEA-3HF to provide a 1.4 M HF concentration) and heated to 65 °C. After 1.5 h, the oligomer is quenched with 1.5 M N ⁇ 4HCO3.
  • the polymer-bound trityl-on oligoribonucleotide is transferred to a 4 mL glass screw top vial and suspended in a solution of 33% ethanolic methylamine/DMSO: 1/1 (0.8 mL) at 65 °C for 15 min.
  • the vial is brought to r.t. TEA « 3HF (0.1 mL) is added and the vial is heated at 65 °C for 15 min.
  • the sample is cooled at -20 °C and then quenched with 1.5 M NH4HCO3.
  • the quenched NH4HCO3 solution is loaded onto a C-18 containing cartridge that had been prewashed with acetonitrile followed by 50 mM TEAA. After washing the loaded cartridge with water, the RNA is detritylated with 0.5% TFA for 13 min. The cartridge is then washed again with water, salt exchanged with 1 M NaCl and washed with water again. The oligonucleotide is then eluted with 30% acetonitrile.
  • Inactive hammerhead ribozymes or binding attenuated control (BAC) oligonucleotides are synthesized by substituting a U for G5 and a U for A14
  • nucleotide substitutions can be introduced in other enzymatic nucleic acid molecules to inactivate the molecule and such molecules can serve as a negative control.
  • the average stepwise coupling yields are typically >98% (Wincott et al, 1995
  • nucleic acid molecules of the present invention can be synthesized separately and joined together post- synthetically, for example by ligation (Moore et al, 1992, Science 256, 9923; Draper et al, Intemational PCT publication No. WO 93/23569; Shabarova et al, 1991, Nucleic Acids Research 19, 4247; Bellon et al, 1997, Nucleosides & Nucleotides, 16, 951; Bellon et al, 1997, Bioconjugate Chem. 8, 204).
  • nucleic acid molecules of the present invention are modified extensively to enhance stability by modification with nuclease resistant groups, for example, 2'- amino, 2'-C-allyl, 2'-flouro, 2'-O-mefhyl, 2'-H (for a review see Usman and Cedergren, 1992, TIBS 17, 34; Usman et al, 1994, Nucleic Acids Symp. Ser. 31, 163).
  • Ribozymes are purified by gel electrophoresis using general methods or are purified by high pressure liquid chromatography (HPLC; See Wincott et al, supra, the totality of which is hereby inco ⁇ orated herein by reference) and are resuspended in water.
  • the ribozyme and antisense constract sequences listed in Tables III to IX may be formed of ribonucleotides or other nucleotides or non- nucleotides. Such ribozymes with enzymatic activity are equivalent to the ribozymes described specifically in the Tables. Optimizing Activity of the nucleic acid molecule of the invention.
  • oligonucleotides are modified to enhance stability and/or enhance biological activity by modification with nuclease resistant groups, for example, 2'-amino, 2'-C-allyl, 2'-flouro, -O- methyl, 2'-H, nucleotide base modifications (for a review see Usman and Cedergren, 1992, TIBS. 17, 34; Usman et al, 1994, Nucleic Acids Symp. Ser. 31, 163; Burgin et al, 1996, Biochemistry , 35, 14090).
  • nuclease resistant groups for example, 2'-amino, 2'-C-allyl, 2'-flouro, -O- methyl, 2'-H, nucleotide base modifications
  • Nucleic acid molecules having chemical modifications which maintain or enhance activity are provided. Such nucleic acid is also generally more resistant to nucleases than unmodified nucleic acid. Thus, in a cell and/or in vivo the activity may not be significantly lowered.
  • Therapeutic nucleic acid molecules delivered exogenously must optimally be stable within cells until translation of the target RNA has been inhibited long enough to reduce the levels of the undesirable protein. This period of time varies between hours to days depending upon the disease state.
  • nucleic acid molecules must be resistant to nucleases in order to function as effective intracellular therapeutic agents. Improvements in the chemical synthesis of RNA and DNA (Wincott et al, 1995 Nucleic Acids Res.
  • nucleic acid-based molecules of the invention will lead to better treatment of the disease progression by affording the possibility of combination therapies (e.g., multiple antisense or enzymatic nucleic acid molecules targeted to different genes, nucleic acid molecules coupled with known small molecule inhibitors, or intermittent treatment with combinations of molecules (including different motifs) and/or other chemical or biological molecules).
  • combination therapies e.g., multiple antisense or enzymatic nucleic acid molecules targeted to different genes, nucleic acid molecules coupled with known small molecule inhibitors, or intermittent treatment with combinations of molecules (including different motifs) and/or other chemical or biological molecules).
  • the treatment of patients with nucleic acid molecules may also include combinations of different types of nucleic acid molecules.
  • Therapeutic nucleic acid molecules e.g., enzymatic nucleic acid molecules and antisense nucleic acid molecules
  • delivered exogenously must optimally be stable within cells until translation of the target RNA has been inhibited long enough to reduce the levels of the undesirable protein.
  • nucleic acid molecules must be resistant to nucleases in order to function as effective intracellular therapeutic agents. Improvements in the chemical synthesis of nucleic acid molecules described in the instant invention and in the art have expanded the ability to modify nucleic acid molecules by introducing nucleotide modifications to enhance their nuclease stability as described above.
  • enhanced enzymatic activity is meant to include activity measured in cells and/or in vivo where the activity is a reflection of both catalytic activity and ribozyme stability.
  • the product of these properties is increased or not significantly (less than 10- fold) decreased in vivo compared to an all RNA ribozyme or all DNA enzyme.
  • nucleic acid catalysts having chemical modifications which maintain or enhance enzymatic activity are provided.
  • Such nucleic acid is also generally more resistant to nucleases than unmodified nucleic acid.
  • the activity may not be significantly lowered.
  • ribozymes are useful in a cell and/or in vivo even if activity over all is reduced 10 fold (Burgin et al, 1996, Biochemistry, 35, 14090).
  • Such ribozymes herein are said to "maintain” the enzymatic activity of an all RNA ribozyme.
  • nucleic acid molecules comprise a 5' and/or a 3'- cap structure.
  • cap stracture is meant chemical modifications, which have been inco ⁇ orated at either terminus of the oligonucleotide (see, for example, Wincott et al, WO 97/26270, inco ⁇ orated by reference herein). These terminal modifications protect the nucleic acid molecule from exonuclease degradation, and may help in delivery and/or localization within a cell.
  • the cap may be present at the 5 '-terminus (5 '-cap) or at the 3 '-terminus (3 '-cap) or may be present on both termini.
  • the 5 '-cap is selected from the group comprising inverted abasic residue (moiety), 4',5'-methylene nucleotide; l-(beta-D-erythrofuranosyl) nucleotide, 4'-thio nucleotide, , carbocyclic nucleotide; 1,5-anhydrohexitol nucleotide; L- nucleotides; alpha-nucleotides; modified base nucleotide; phosphorodithioate linkage; t/zreo-pentofuranosyl nucleotide; acyclic 3',4'-seco nucleotide; acyclic 3,4- dihydroxybutyl nucleotide; acyclic 3,5-dihydroxypentyl nucleotide, 3'-3 '-inverted nucleotide moiety; 3 '-3 '-inverted abasic moiety; 3'-2'
  • the 3 '-cap is selected from a group comprising, 4',5'-mefhylene nucleotide; l-(beta-D-erythrofuranosyl) nucleotide; 4'- thio nucleotide, carbocyclic nucleotide; 5'-amino-alkyl phosphate; l,3-diamino-2- propyl phosphate, 3-aminopropyl phosphate; 6-aminohexyl phosphate; 1,2- aminododecyl phosphate; hydroxypropyl phosphate; 1,5-anhydrohexitol nucleotide;
  • non-nucleotide any group or compound which can be inco ⁇ orated into a nucleic acid chain in the place of one or more nucleotide units, including either sugar and/or phosphate substitutions, and allows the remaining bases to exhibit their enzymatic activity.
  • the group or compound is abasic in that it does not contain a commonly recognized nucleotide base, such as adenosine, guanine, cytosine, uracil or thymine.
  • alkyl refers to a saturated aliphatic hydrocarbon, including straight-chain, branched-chain, and cyclic alkyl groups.
  • the alkyl group has 1 to 12 carbons. More preferably it is a lower alkyl of from 1 to 7 carbons, more preferably 1 to 4 carbons.
  • the term also includes alkenyl groups which are unsaturated hydrocarbon groups containing at least one carbon-carbon double bond, including straight-chain, branched-chain, and cyclic groups.
  • the alkenyl group has 1 to 12 carbons. More preferably it is a lower alkenyl of from 1 to 7 carbons, more preferably 1 to 4 carbons.
  • alkyl also includes alkynyl groups which have an unsaturated hydrocarbon group containing at least one carbon-carbon triple bond, including straight-chain, branched-chain, and cyclic groups.
  • the alkynyl group has 1 to 12 carbons. More preferably it is a lower alkynyl of from 1 to 7 carbons, more preferably 1 to 4 carbons.
  • alkyl groups may also include aryl, alkylaryl, carbocyclic aryl, heterocyclic aryl, amide and ester groups.
  • An "aryl” group refers to an aromatic group which has at least one ring having a conjugated ⁇ electron system and includes carbocyclic aryl, heterocyclic aryl and biaryl groups, all of which may be optionally substituted.
  • the preferred substituent(s) of aryl groups are halogen, trihalomethyl, hydroxyl, SH, OH, cyano, alkoxy, alkyl, alkenyl, alkynyl, and amino groups.
  • alkylaryl refers to an alkyl group (as described above) covalently joined to an aryl group (as described above).
  • Carbocyclic aryl groups are groups wherein the ring atoms on the aromatic ring are all carbon atoms. The carbon atoms are optionally substituted.
  • Heterocyclic aryl groups are groups having from 1 to 3 heteroatoms as ring atoms in the aromatic ring and the remainder of the ring atoms are carbon atoms.
  • Suitable heteroatoms include oxygen, sulfur, and nitrogen, and include furanyl, thienyl, pyridyl, pyrrolyl, N-lower alkyl pyrrolo, pyrimidyl, pyrazinyl, imidazolyl and the like, all optionally substituted.
  • An "amide” refers to an -C(O)-NH-R,. where R is either alkyl, aryl, alkylaryl or hydrogen.
  • An “ester” refers to an -C(O)-OR', where R is either alkyl, aryl, alkylaryl or hydrogen.
  • nucleotide as used herein is as recognized in the art to include natural bases (standard), and modified bases well known in the art. Such bases are generally located at the 1' position of a nucleotide sugar moiety. Nucleotides generally comprise a base, sugar and a phosphate group. The nucleotides can be unmodified or modified at the sugar, phosphate and/or base moiety, (also referred to interchangeably as nucleotide analogs, modified nucleotides, non-natural nucleotides, non-standard nucleotides and other; see for example, Usman and McSwiggen, supra; Eckstein et al, International PCT Publication No.
  • base modifications that can be introduced into nucleic acid molecules include, inosine, purine, pyridin-4-one, pyridin-2-one, phenyl, pseudouracil, 2, 4, 6-trimethoxy benzene, 3 -methyl uracil, dihydrouridine, naphthyl, aminophenyl, 5-alkylcytidines (e.g., 5-methylcytidine), 5-alkyluridines (e.g., ribothymidine), 5-halouridine (e.g., 5-bromouridine) or 6-azapyrimidines or 6- alkylpyrimidines (e.g.
  • modified bases in this aspect is meant nucleotide bases other than adenine, guanine, cytosine and uracil at 1' position or their equivalents; such bases may be used at any position, for example, within the catalytic core of an enzymatic nucleic acid molecule and/or in the substrate-binding regions of the nucleic acid molecule.
  • the invention features modified ribozymes with phosphate backbone modifications comprising one or more phosphorothioate, phosphorodithioate, methylphosphonate, mo ⁇ holino, amidate carbamate, carboxymethyl, acetamidate, polyamide, sulfonate, sulfonamide, sulfamate, formacetal, thioformacetal, and/or alkylsilyl, substitutions.
  • abasic sugar moieties lacking a base or having other chemical groups in place of a base at the 1' position, (for more details, see Wincott et al, Intemational PCT publication No. WO 97/26270).
  • unmodified nucleoside is meant one of the bases adenine, cytosine, guanine, thymine, uracil joined to the 1' carbon of ⁇ -D-ribo-furanose.
  • modified nucleoside any nucleotide base which contains a modification in the chemical structure of an unmodified nucleotide base, sugar and/or phosphate.
  • amino 2'-NH 2 or 2'-O- NH , which may be modified or unmodified.
  • modified groups are described, for example, in Eckstein et al,
  • nucleic acid molecules may also include combinations of different types of nucleic acid molecules.
  • therapies may be devised which include a mixture of ribozymes (including different ribozyme motifs), antisense and/or 2-5A chimera molecules to one or more targets to alleviate symptoms of a disease.
  • nucleic acid molecules Methods for the delivery of nucleic acid molecules are described in Akhtar et al, 1992, Trends Cell Bio., 2, 139; and Delivery Strategies for Antisense Oligonucleotide Therapeutics, ed. Akhtar, 1995 which are both inco ⁇ orated herein by reference.
  • Sullivan et al, PCT WO 94/02595 further describes the general methods for delivery of enzymatic RNA molecules. These protocols may be utilized for the delivery of virtually any nucleic acid molecule.
  • Nucleic acid molecules may be administered to cells by a variety of methods known to those familiar to the art, including, but not restricted to, encapsulation in liposomes, by iontophoresis, or by inco ⁇ oration into other vehicle ' s " such as hydrogels, cyclodextrins, biodegradable nanocapsules, and bioadhesive microspheres.
  • nucleic acid molecules may be directly delivered ex vivo to cells or tissues with or without the aforementioned vehicles.
  • the nucleic acid/vehicle combination is locally delivered by direct injection or by use of a catheter, infusion pump or stent.
  • nucleic acid delivery and administration are provided in Sullivan et al, supra, Draper et al, PCT WO93/23569, Beigelman et al, PCT WO99/05094, and Klimuk et al, PCT WO99/04819 all of which have been inco ⁇ orated by reference herein.
  • nucleic acid molecules of the instant invention used to treat pulmonary diseases and disorders, may be administered directly to the lungs via pulmonary delivery.
  • the molecules of the instant invention can be used as pharmaceutical agents.
  • Pharmaceutical agents prevent, inhibit the occurrence, or treat (alleviate a symptom to some extent, preferably all of the symptoms) of a disease state in a patient.
  • the negatively charged polynucleotides of the invention can be administered (e.g., RNA, DNA or protein) and introduced into a patient by any standard means, with or without stabilizers, buffers, and the like, to form a pharmaceutical composition.
  • RNA, DNA or protein e.g., RNA, DNA or protein
  • standard protocols for formation of liposomes can be followed.
  • the compositions of the present invention may also be formulated and used as tablets, capsules or elixirs for oral administration; suppositories for rectal administration; sterile solutions; suspensions for injectable administration; and other compositions known in the art.
  • the present invention also includes pharmaceutically acceptable formulations of the compounds described.
  • formulations include salts of the above compounds, e.g., acid addition salts, including salts of hydrochloric, hydrobromic, acetic acid, and benzene sulfonic acid.
  • a pharmacological composition or formulation refers to a composition or formulation in a form suitable for administration, e.g., systemic administration, into a cell or patient, preferably a human. Suitable forms, in part, depend upon the use or the route of entry, for example oral, transdermal, or by injection. Such forms should not prevent the composition or formulation from reaching a target cell (i.e., a cell to which the negatively charged polymer is desired to be delivered to). For example, pharmacological compositions injected into the blood stream should be soluble.
  • systemic administration is meant in vivo systemic abso ⁇ tion or accumulation of drags in the blood stream followed by distribution throughout the entire body.
  • Administration routes that lead to systemic abso ⁇ tion include, without limitations: intravenous, subcutaneous, intraperitoneal, inhalation, oral, intrapulmonary and intramuscular.
  • Each of these administration routes exposes the desired negatively charged polymers, e.g., nucleic acids, to an accessible diseased tissue.
  • the rate of entry of a drug into the circulation has been shown to be a function of molecular weight or size.
  • the use of a liposome or other drug carrier comprising the compounds of the instant invention can potentially localize the drag, for example, in certain tissue types, such as the tissues of the reticular endothelial system (RES).
  • RES reticular endothelial system
  • a liposome formulation that can facilitate the association of drug with the surface of cells, such as, lymphocytes and macrophages is also useful. This approach may provide enhanced delivery of the drug to target cells by taking advantage of the specificity of macrophage and lymphocyte immune recognition of abnormal cells, such as cancer cells.
  • compositions or formulation that allows for the effective distribution of the nucleic acid molecules of the instant invention in the physical location most suitable for their desired activity.
  • agents suitable for formulation with the nucleic acid molecules of the instant invention include: P-glycoprotein inhibitors (such as Pluronic P85) which can enhance entry of drags into the CNS (Jolliet-Riant and Tillement, 1999, Fundam. Clin. Pharmacol, 13, 16-26); biodegradable polymers, such as poly (DL-lactide-coglycolide) microspheres for sustained release delivery after intracerebral implantation (Emerich, DF et al, 1999, Cell Transplant, 8, 47-58) Alkermes, Inc.
  • nanoparticles such as those made of polybutylcyanoacrylate, which can deliver drags across the blood brain barrier and can alter neuronal uptake mechanisms (Prog Neuropsychopharmacol Biol Psychiatry, 23, 941-949, 1999).
  • Other non-limiting examples of delivery strategies for the nucleic acid molecules of the instant invention include material described in Boado et al, 1998, J. Pharm. Sci., 87, 1308-1315; Tyler et al, 1999, FEBS Lett, 421, 280-284; Pardridge et al, 1995, PNAS USA., 92, 5592-5596; Boado, 1995, Adv. Drug Delivery Rev., 15, 73-107; Aldrian-Herrada et al, 1998, Nucleic Acids Res., 26, 4910-4916; and Tyler et al, 1999, PNAS USA., 96, 7053-7058.
  • the invention also features the use of the composition comprising surface- modified liposomes containing poly (ethylene glycol) lipids (PEG-modified, or long- circulating liposomes or stealth liposomes).
  • PEG-modified, or long- circulating liposomes or stealth liposomes These formulations offer a method for increasing the accumulation of drags in target tissues.
  • This class of drag carriers resists opsonization and elimination by the mononuclear phagocytic system (MPS or RES), thereby enabling longer blood circulation times and enhanced tissue exposure for the encapsulated drag (Lasic et al. Chem. Rev. 1995, 95, 2601-2627; Ishiwata et al, Chem. Pharm. Bull 1995, 43, 1005-1011). All inco ⁇ orated by reference herein.
  • liposomes have been shown to accumulate selectively in tumors, presumably by extravasation and capture in the neovascularized target tissues (Lasic et al, Science 1995, 267, 1275-1276; Oku et al, 1995, Biochim. Biophys. Acta, 1238, 86- 90). All inco ⁇ orated by reference herein.
  • the long-circulating liposomes enhance the pharmacokinetics and pharmacodynamics of DNA and RNA, particularly compared to conventional cationic liposomes which are known to accumulate in tissues of the MPS (Liu et al, J. Biol. Chem. 1995, 42, 24864-24870; Choi et al, International PCT Publication No.
  • WO 96/10391 Ansell et al, International PCT Publication No. WO 96/10390; Holland et al, Intemational PCT Publication No. WO 96/10392; all of which are inco ⁇ orated by reference herein).
  • Long-circulating liposomes are also likely to protect drags from nuclease degradation to a greater extent compared to cationic liposomes, based on their ability to avoid accumulation in metabolically aggressive MPS tissues such as the liver and spleen.
  • the invention features the use of methods to deliver the nucleic acid molecules of the instant invention to hematopoietic cells, including monocytes and lymphocytes. These methods are described in detail by Hartmann et al, 1998, J. Phamacol Exp. Ther., 285(2), 920-928; Kronenwett et al, 1998, Blood, 91(3), 852- 862; Filion and Phillips, 1997, Biochim. Biophys. Acta., 1329(2), 345-356; Ma and Wei, 1996, Leuk. Res., 20(11/12), 925-930; and Bongartz et al, 1994, Nucleic Acids Research, 22(22), 4681-8.
  • Such methods include the use of free oligonucleotide, cationic lipid formulations, liposome formulations including pH sensitive liposomes and immunoliposomes, and bioconjugates including oligonucleotides conjugated to fusogenic peptides, for the transfection of hematopoietic cells with oligonucleotides.
  • compositions prepared for storage or administration which include a pharmaceutically effective amount of the desired compounds in a pharmaceutically acceptable carrier or diluent.
  • Acceptable carriers or diluents for therapeutic use are well known in the pharmaceutical art, and are described, for example, in Remington 's Pharmaceutical Sciences, Mack Publishing Co. (A.R. Gennaro edit. 1985) hereby inco ⁇ orated by reference herein.
  • preservatives, stabilizers, dyes and flavoring agents may be provided. These include sodium benzoate, sorbic acid and esters of j9-hydroxybenzoic acid.
  • antioxidants and suspending agents may be used.
  • a pharmaceutically effective dose is that dose required to prevent, inhibit the occurrence, or treat (alleviate a symptom to some extent, preferably all of the symptoms) of a disease state.
  • the pharmaceutically effective dose depends on the type of disease, the composition used, the route of administration, the type of mammal being treated, the physical characteristics of the specific mammal under consideration, concurrent medication, and other factors which those skilled in the medical arts will recognize. Generally, an amount between 0.1 mg/kg and 100 mg/kg body weight/day of active ingredients is administered dependent upon potency of the negatively charged polymer.
  • the nucleic acid molecules of the present invention may also be administered to a patient in combination with other therapeutic compounds to increase the overall therapeutic effect.
  • the use of multiple compounds to treat an indication may increase the beneficial effects while reducing the presence of side effects.
  • Oxygen therapy, bronchodilators, corticosteroids, antibacterials, vaccinations, acetylcysteine, mucokinetic agents, and DNase (Pulmozyme) are non-limiting examples of compounds and/or methods that can be combined with or used in conjunction with the nucleic acid molecules (e.g. ribozymes and antisense molecules) of the instant invention.
  • ribozymes and antisense molecules e.g. ribozymes and antisense molecules
  • nucleic acid molecules of the instant invention can be expressed within cells from eukaryotic promoters (e.g., Izant and Weintraub, 1985, Science, 229, 345; McGarry and Lindquist, 1986, Proc. Natl Acad. Sci., USA 83, 399; Scanlon et al, 1991, Proc. Natl. Acad. Sci. USA, 88, 10591-5; Kashani- Sabet et al, 1992, Antisense Res. Dev., 2, 3-15; Dropulic et al, 1992, J. Virol, 66, 1432-41; Weerasinghe et al, 1991, J.
  • eukaryotic promoters e.g., Izant and Weintraub, 1985, Science, 229, 345; McGarry and Lindquist, 1986, Proc. Natl Acad. Sci., USA 83, 399; Scanlon et al, 1991, Proc.
  • nucleic acids can be augmented by their release from the primary transcript by a ribozyme (Draper et al, PCT WO 93/23569, and Sullivan et al, PCT WO 94/02595; Ohkawa et al, 1992, Nucleic Acids Symp. Ser., 27, 15-6; Taira et ⁇ /., 1991, Nucleic Acids Res., 19, 5125- 30; Ventura et al, 1993, Nucleic Acids Res., 21, 3249-55; Chowrira et al, 1994, J. Biol. Chem., 269, 25856; all of these references are hereby inco ⁇ orated in their totalities by reference herein).
  • a ribozyme Draper et al, PCT WO 93/23569, and Sullivan et al, PCT 94/02595; Ohkawa et al, 1992, Nucleic Acids Symp. Ser., 27, 15-6
  • RNA molecules of the present invention are preferably expressed from transcription units (see, for example, Couture et al, 1996, TIG., 12, 510) inserted into DNA or RNA vectors.
  • the recombinant vectors are preferably DNA plasmids or viral vectors. Ribozyme expressing viral vectors could be constructed based on, but not limited to, adeno-associated virus, retrovirus, adenovirus, or alphaviras.
  • the recombinant vectors capable of expressing the nucleic acid molecules are delivered as described above, and persist in target cells.
  • viral vectors may be used that provide for transient expression of nucleic acid molecules. Such vectors might be repeatedly administered as necessary.
  • nucleic acid molecule binds to the target mRNA.
  • Delivery of nucleic acid molecule expressing vectors could be systemic, such as by intravenous or intra-muscular administration, by administration to target cells ex- planted from the patient followed by reintroduction into the patient, or by any other means that would allow for introduction into the desired target cell (for a review, see Couture et al, 1996, TIG., 12, 510).
  • the invention features an expression vector comprising a nucleic acid sequence encoding at least one of the nucleic acid molecules disclosed in the instant invention.
  • the nucleic acid sequence encoding the nucleic acid molecule of the instant invention is operably linked in a manner which allows expression of that nucleic acid molecule.
  • the invention features an expression vector comprising: a) a transcription initiation region (e.g., eukaryotic pol I, II or in initiation region); b) a transcription termination region (e.g., eukaryotic pol I, ⁇ or HI termination region); c) a nucleic acid sequence encoding at least one of the nucleic acid catalyst of the instant invention; and wherein said sequence is operably linked to said initiation region and said termination region, in a manner which allows expression and/or delivery of said nucleic acid molecule.
  • the vector may optionally include an open reading frame (ORF) for a protein operably linked on the 5' side or the 3'-side of the sequence encoding the nucleic acid catalyst of the invention; and/or an intron (intervening sequences).
  • ORF open reading frame
  • RNA polymerase I eukaryotic RNA polymerase I
  • poly TL RNA polymerase II
  • RNA polymerase HI RNA polymerase HI
  • Transcripts from pol H or pol HI promoters will be expressed at high levels in all cells; the levels of a given pol H promoter in a given cell type will depend on the nature of the gene regulatory sequences (enhancers, silencers, etc.) present nearby.
  • Prokaryotic RNA polymerase promoters are also used, providing that the prokaryotic RNA polymerase enzyme is expressed in the appropriate cells (Elroy-Stein and Moss, 1990, Proc. Natl. Acad. Sci.
  • nucleic acid molecules such as ribozymes expressed from such promoters can function in mammalian cells (e.g. Kashani-Sabet et al, 1992, Antisense Res. Dev., 2, 3-15; Ojwang et al, 1992, Proc. Nat Acad. Sci. USA, 89, 10802-6; Chen etal, 1992, Nucleic Acids Res., 20, 4581-9; Yu et al, 1993, Proc. Natl. Acad. Sci. USA, 90, 6340-4; L'Huillier et al, 1992, EMBO J., 11, 4411-8; Lisziewicz et al, 1993, Proc. Natl. Acad.
  • transcription units such as the ones derived from genes encoding U6 small nuclear (snRNA), transfer RNA (tRNA) and adenovirus VA RNA are useful in generating high concentrations of desired RNA molecules such as ribozymes in cells (Thompson et al, supra; Couture and Stinchcomb, 1996, supra; Noonberg et al, 1994, Nucleic Acid Res., 22, 2830; Noonberg et al, US Patent No.
  • ribozyme transcription units can be inco ⁇ orated into a variety of vectors for introduction into mammalian cells, including but not restricted to, plasmid DNA vectors, viral DNA vectors (such as adenovirus or adeno-associated virus vectors), or viral RNA vectors (such as retroviral or alphaviras vectors) (for a review, see Couture and Stinchcomb, 1996, supra).
  • plasmid DNA vectors such as adenovirus or adeno-associated virus vectors
  • viral RNA vectors such as retroviral or alphaviras vectors
  • the invention features an expression vector comprising a nucleic acid sequence encoding at least one of the nucleic acid molecules of the invention, in a manner which allows expression of that nucleic acid molecule.
  • the expression vector comprises in one embodiment; a) a transcription initiation region; b) a transcription termination region; c) a nucleic acid sequence encoding at least one said nucleic acid molecule; and wherein said sequence is operably linked to said initiation region and said termination region, in a manner which allows expression and/or delivery of said nucleic acid molecule.
  • the expression vector comprises: a) a transcription initiation region; b) a transcription termination region; c) an open reading frame; d) a nucleic acid sequence encoding at least one said nucleic acid molecule, wherein said sequence is operably linked to the 3 '-end of said open reading frame; and wherein said sequence is operably linked to said initiation region, said open reading frame and said termination region, in a manner which allows expression and/or delivery of said nucleic acid molecule.
  • the expression vector comprises: a) a transcription initiation region; b) a transcription termination region; c) an intron; d) a nucleic acid sequence encoding at least one said nucleic acid molecule; and wherein said sequence is operably linked to said initiation region, said intron and said termination region, in a manner which allows expression and or delivery of said nucleic acid molecule.
  • the expression vector comprises: a) a transcription initiation region; b) a transcription termination region; c) an intron; d) an open reading frame; e) a nucleic acid sequence encoding at least one said nucleic acid molecule, wherein said sequence is operably linked to the 3'-end of said open reading frame; and wherein said sequence is operably linked to said initiation region, said intron, said open reading frame and said termination region, in a manner which allows expression and/or delivery of said nucleic acid molecule.
  • the reporter system consists of a plasmid construct, termed pMUC5AC- EGFP, bearing a gene coding for Green Fluorescent Protein (GFP).
  • GFP Green Fluorescent Protein
  • the promoter region of the GFP gene is replaced by a portion of the Mucin 5AC promoter sufficient to direct efficient transcription of the GFP gene.
  • the plasmid also contains the neomycin drug resistance gene.
  • NCI-H292 (ATCC CRL-1848), is derived from a human lung mucoepidermoid carcinoma.
  • the cells retain mucoepidermoid characteristics in culture and endogenously express mucin 5AC and mucin 2.
  • the ⁇ MUC5AC-EGFP plasmid was transfected into NCI-H292 using a cationic lipid formulation. Following transfection, the cells were subjected to limiting dilution cloning under selection by 600 ⁇ g/mL Geneticin.
  • a clonal cell line highly responsive to mucin promoter induction designated
  • H292/MUC5AC/EGFP Clone8 (H292 Clone 8) was selected as the reporter line for subsequent studies.
  • the process for Target Discovery is described in Jarvis et al, International PCT publication No. WO 98/50530, inco ⁇ orated by reference herein in its entirety including the Figures.
  • a ribozyme library was constracted with oligonucletides containing ribozymes with two randomized regions comprising six-nucleotide binding "arms" (Stem I and Stem HI of a ribozyme-substrate complex). Oligo sequence 5' and 3' of the ribozyme contains restriction endonuclease cleavage sites for cloning. The 3' trailing sequence forms a stem-loop for priming DNA polymerase extension to form a double stranded molecule.
  • the double-stranded ribozyme library was cloned into the U6+27 transcription unit located in the 5' LTR region of a retroviral vector containing the human nerve growth factor receptor (hNGFr) reporter gene. Positioning the U6+27/ribozyme transcription unit in the 5' LTR results in a duplication of the transcription unit when the vector integrates into the host cell genome. As a result, the ribozyme is transcribed by RNA polymerase HI from U6+27 and by RNA polymerase H activity directed by the 5' LTR. The ribozyme library was packaged into retroviral particles that were used to infect and transduce H292 Clone 8 cells.
  • hNGFr human nerve growth factor receptor
  • FIG. 5A and 5B describe the generalized scheme used in the ribozyme library constraction and target discovery.
  • randomized region is meant a region of completely random sequence and/or partially random sequence.
  • completely random sequence is meant a sequence wherein theoretically there is equal representation of A, T, G and C nucleotides or modified derivatives thereof, at each position in the sequence.
  • partially random sequence is meant a sequence wherein there is an unequal representation of A, T, G and C nucleotides or modified derivatives thereof, at each position in the sequence.
  • a partially random sequence can therefore have one or more positions of complete randomness and one or more positions with defined nucleotides.
  • Sorting of ribozyme library-containing cells was performed to enrich for cells that produce less GFP after treatment with PCM and PMA.
  • Lower GFP production may be due to ribozyme action upon genes involved in the activation of the mucin promoter.
  • ribozymes may directly target the mucin/GFP transcript resulting in reduced GFP expression.
  • Cells were seeded at a density of 1 x 10 per 150 cm style cell culture flasks. After 72 hours the standard cell culture medium was replaced with medium without fetal bovine serum. After 24 hours of seram deprivation the cells were treated with serum-containing medium supplemented with PCM (to 40%) and PMA (to 50 nM) to induced GFP production via the mucin promoter. After 20 to 22 hours, cells were monitored for GFP level on a FACStar Plus cell sorter.
  • Sorting was performed if 90% of ribozyme library cells from an unsorted control sample were induced to produce GFP above background levels. Two cell fractions were collected in each round of sorting.
  • the Ml gate collected cells in luminescence channels 1 to 4.5; those cells with the lowest GFP signal (5% of the induced population).
  • the M2 sort gate collected cells in luminescence channels 4.5 to 20; cells with low GFP signal (10% of the induced population).
  • the Ml and M2 fractions together represented the 15% of the induced population responding least to the GFP induction treatment.
  • the Ml and M2 fractions wee cultured separately and representative portions of each were cryopreserved after each round of sorting.
  • the Ml fraction Prior to the third round of sorting the Ml fraction showed a three-fold enrichment of GFP negative cells. Prior to the fourth round of sorting both the Ml and M2 fractions were significantly enriched in cells unresponsive to the GFP induction treatment.
  • the Ml fraction was selected to generate a database of ribozymes present in the sorted cells.
  • Genomic DNA was obtained from sorted ribozyme library cells by standard methods. Nested polymerase chain reaction (PCR) primers (Sequence ID Nos. 5468 and 5469) that hybridized to the retroviral vector 5' and 3' of the ribozyme were used to recover and amplify the ribozyme sequences from the Clone 8 library cell DNA. The PCR product was ligated into a bacterial cloning vector. Two methods were developed to use the recovered ribozyme library, in plasmid form, to generate a database of ribozyme binding arm sequences. In the first approach the library was cloned into E. coli. DNA was prepared by plasmid isolation from bacterial colonies or by direct colony.
  • PCR polymerase chain reaction
  • PCR and _ribozyme..arm sequence. was determined. Over 450 sequences have been obtained by this method.
  • a second method used the ribozyme library to transfect H292 Clone 8 cells. Clonal lines of stably transfected cells were established and induced with PCM and PMA. Those lines which failed to respond to GFP induction were probed by PCR for single ribozyme integration events. Over 300 sequences were obtained in this manner. The unique ribozyme sequences obtained by both methods were added to a Target Sequence Tag (TST) database.
  • TST Target Sequence Tag
  • Ribozyme binding arm sequences were compared to public and private gene data banks. Gene matches were compiled according to perfect and imperfect matches. Potential gene targets were categorized by the number of different ribozyme sequences matching each gene. Multiple ribozyme matches have been found for 180 genes. Genes with more than one perfect ribozyme match were given close attention. A total of 34 genes have been verified to date to have multiple perfect ribozyme matches. Of those at least 17 have protein products of known function.
  • hCLCAl as a candidate for target validation was based on bioinformatics and on emerging data in murine models of mucous hypersecretion in the trachea and lung.
  • Two ribozymes (Seq. ID Nos. 2332 and 2273) recovered from cells that no longer respond to mucin promoter/GFP induction match perfectly to hCLCAl.
  • a third has a single mismatch.
  • Evidence from two murine models indicates a correlation between mucous hypersecretion in the lung and strong upregulation of gob-5 (GenBank ABO17156), a murine homologue of hCLCAl.
  • GeneBIoc reagents were designed (Table IX) to the hCLCAl cDNA sequence (GenBank AF039400). GeneBIoc reagents are complexed with a cationic lipid formulation prior to administration to H292/MUC5AC/GFP Clone 8 cells. Concentrations of the GeneBIoc reagents administered range from 30 nM to 120 nM at cationic lipid concentrations of 4-6 ⁇ g/mL. Cells are treated with GeneBIoc reagents for 72 to 96 hours.
  • the sequence of human CLCAl is screened for accessible sites using a computer-folding algorithm. Regions of the RNA are identified that do not form secondary folding structures. These regions contain potential ribozyme and/or antisense binding/cleavage sites. The sequences of these binding/cleavage sites are shown in Tables III-IX.
  • Ribozyme target sites are chosen by analyzing sequences of Human CLCAl (GenBank accession numbers: NM_001285 and AF039400) and prioritizing the sites on the basis of folding. Ribozymes are designed that could bind each target and are individually analyzed by computer folding (Christoffersen et al, 1994 J. Mol. Struc. Theochem, 311, 273; Jaeger et al, 1989, Proc. Natl. Acad. Sci. USA, 86, 7706) to assess whether the ribozyme sequences fold into the appropriate secondary structure. Those ribozymes with unfavorable intramolecular interactions between the binding arms and the catalytic core are eliminated from consideration. As noted below, varying . binding arm lengths can be chosen to optimize activity. Generally, at least 5 bases on each arm are able to bind to, or otherwise interact with, the target RNA.
  • Example 11 Chemical Synthesis and Purification of Ribozymes and Antisense for Efficient Cleavage and/or blocking of CLCAl RNA
  • Ribozymes and antisense constructs are designed to anneal to various sites in the RNA message.
  • the binding arms of the ribozymes are complementary to the target site sequences described above, while the antisense constructs are fully complimentary to the target site sequences described above.
  • the ribozymes and antisense constructs were chemically synthesized. The method of synthesis used followed the procedure for normal RNA synthesis as described above and in Usman et al, (1987 J. Am. Chem.
  • Ribozymes and antisense constructs are purified by gel electrophoresis using general methods or are purified by high pressure liquid chromatography (HPLC; see Wincott et al, supra; the totality of which is hereby inco ⁇ orated herein by reference) and are resuspended in water.
  • HPLC high pressure liquid chromatography
  • the sequences of the chemically synthesized ribozymes and antisense constructs used in this study are shown below in Table III-IX.
  • Particular conditions and disease states that can be associated with CLCAl expression modulation include but are not limited to Chronic Obstructive Pulmonary Disease (COPD), chronic bronchitis, asthma, cystic fibrosis, obstructive bowel syndrome, and any other diseases or conditions that are related to or will respond to the levels of CLCAl in a cell or tissue, alone or in combination with other therapies.
  • COPD Chronic Obstructive Pulmonary Disease
  • chronic bronchitis chronic bronchitis
  • asthma chronic bronchitis
  • cystic fibrosis obstructive bowel syndrome
  • any other diseases or conditions that are related to or will respond to the levels of CLCAl in a cell or tissue alone or in combination with other therapies.
  • the nucleic acid molecules of the present invention may also be administered to a patient in combination with other therapeutic compounds to increase the overall therapeutic effect.
  • the use of multiple compounds to treat an indication may increase the beneficial effects while reducing the presence of side effects.
  • Oxygen therapy, bronchodilators, corticosteroids, antibacterials, vaccinations, acetylcysteine, mucokinetic agents, and DNase (Pulmozyme) are non-limiting examples of methods and/or treatments that can be used in combination with nucleic acid molecules of the invention.
  • Those skilled in the art will recognize that other drug compounds and therapies can be similarly and readily combined with the nucleic acid molecules of the instant invention (e.g. ribozymes and antisense molecules) and are, therefore, within the scope of the instant invention.
  • Example 8 The cell culture system described in Example 8 can be used to evaluate nucleic acid molecules of the invention for efficacy in CLCAl and mucin modulation. Animal Models
  • COPD chronic pulmonary disease
  • cystic fibrosis animal models relevant to disease states such as COPD and cystic fibrosis. These models can be used to determine efficacy of the nucleic acid molecules of the instant invention targeting such disease states or conditions.
  • Animal models for chronic pulmonary disease (COPD) are described by Shapiro, 2000, Am. J. Respir. Cell Mol Biol, 22(1), 4-7; Hogg, 1998, Ika Daigaku Zasshi, 56(3), 429-432; and Garssen et al, 1997, Inhalation Toxicol, 9(6), 581-599.
  • cystic fibrosis are described by Kent et al, 1997, J. Clin.
  • the nucleic acid molecules of this invention may be used as diagnostic tools to examine genetic drift and mutations within diseased cells or to detect the presence of CLCAl RNA in a cell.
  • the close relationship between ribozyme activity and the structure of the target RNA allows the detection of mutations in any region of the molecule which alters the base-pairing and three- dimensional stracture of the target RNA.
  • ribozymes described in this invention one may map nucleotide changes which are important to RNA structure and function in vitro, as well as in cells and tissues. Cleavage of target RNAs with ribozymes may be used to inhibit gene expression and define the role (essentially) of specified gene products in the progression of disease.
  • ribozymes of this invention include detection of the presence of mRNAs associated with CLCAl -related condition. Such RNA is detected by determining the presence of a cleavage product after treatment with a ribozyme using standard methodology.
  • ribozymes which can cleave only wild-type or mutant forms of the target RNA are used for the assay.
  • the first ribozyme is used to identify wild-type RNA present in the sample and the second ribozyme will be used to identify mutant RNA in the sample.
  • synthetic substrates of both wild-type and mutant RNA will be cleaved by both ribozymes to demonstrate the relative ribozyme efficiencies in the reactions and the absence of cleavage of the "non-targeted" RNA species.
  • the cleavage products from the synthetic substrates will also serve to generate size markers for the analysis of wild-type and mutant RNAs in the sample population.
  • each analysis can require two ribozymes, two substrates and one unknown sample, which will be combined into six reactions.
  • the presence of cleavage products will be determined using an RNAse protection assay so that full-length and cleavage fragments of each RNA can be analyzed in one lane of a polyacrylamide gel. It is not absolutely required to quantify the results to gain insight into the expression of mutant RNAs and putative risk of the desired phenotypic changes in target cells.
  • the expression of mRNA whose protein product is implicated in the development of the phenotype i.e., CLCAl
  • a qualitative comparison of RNA levels will be adequate and will decrease the cost of the initial diagnosis. Higher mutant form to wild-type ratios will be correlated with higher risk whether RNA levels are compared qualitatively or quantitatively.
  • sequence-specific enzymatic nucleic acid molecules of the instant invention might have many of the same applications for the study of RNA that DNA restriction endonucleases have for the study of DNA (Nathans et al, 1975 Ann. Rev. Biochem. 44:273).
  • the pattern of restriction fragments could be used to establish sequence relationships between two related RNAs, and large RNAs could be specifically cleaved to fragments of a size more useful for study.
  • the ability to engineer sequence specificity of the enzymatic nucleic acid molecule is ideal for cleavage of RNAs of unknown sequence.
  • Applicant describes the use of nucleic acid molecules to down-regulate gene expression of target genes in bacterial, microbial, fungal, viral, and eukaryotic systems including plant, or mammalian cells.
  • Reaction mechanism attack by the 3' -OH of guanosine to generate cleavage products with 3'-OH and 5'-guanosine.
  • the small (4-6 nt) binding site may make this ribozyme too non-specific for targeted RNA cleavage, however, the Tetrahymena group I intron has been used to repair a "defective" D -galactosidase message by the ligation of new D- galactosidase sequences onto the defective message [ xu ].
  • RNAse P RNA Ml RNA
  • RNA portion of a ubiquitous ribonucleoprotein enzyme • RNA portion of a ubiquitous ribonucleoprotein enzyme.
  • Reaction mechanism possible attack by M -OH to generate cleavage products with 3' -OH and 5'-phosphate.
  • RNAse P is found throughout the prokaryotes and eukaryotes.
  • the RNA subunit has been sequenced from bacteria, yeast, rodents, and primates.
  • Reaction mechanism 2' -OH of an internal adenosine generates cleavage products with 3' -OH and a "lariat" RNA containing a 3'-5' and a 2'-5' branch point.
  • Reaction mechanism attack by 2'-OH 5' to the scissile bond to generate cleavage products with 2',3'-cyclic phosphate and 5'-OH ends.
  • Reaction mechanism attack by 2'-OH 5' to the scissile bond to generate cleavage products with 2',3'-cyclic phosphate and 5'-OH ends.
  • Reaction mechanism attack by 2'-OH 5' to the scissile bond to generate cleavage products with 2',3' -cyclic phosphate and 5' -OH ends. • 3 known members of this class. Found in three plant pathogen (satellite RNAs of the tobacco ringspot virus, arabis mosaic virus and chicory yellow mottle virus) which uses RNA as the infectious agent.
  • Ligation activity (in addition to cleavage activity) makes ribozyme amenable to engineering through in vitro selection [ x xv ]
  • HDV Hepatitis Delta Virus
  • Folded ribozyme contains a pseudoknot structure [ x1 ].
  • Reaction mechanism attack by 2'-OH 5' to the scissile bond to generate cleavage products with 2',3'-cyclic phosphate and 5' -OH ends.
  • Circular form of HDV is active and shows increased nuclease stability [ xli ]
  • RNA is a catalytic component of a DNA endonuclease involved in intron mobility.
  • Cell Cell (Cambridge, Mass.) (1995), 83(4), 529-38.
  • xxi Griffin, Edmund A., Jr.; Qin, Zhifeng; Michels, Williams J., Jr.; Pyle, Anna Marie.
  • Group II intron ribozymes that cleave DNA and RNA linkages with similar efficiency, and lack contacts with substrate 2'-hydroxyl groups. Chem. Biol. (1995), 2(11), 761-70. x ⁇ . Michel, Francois; Ferat, Jean Luc. Structure and activities of group II introns. Annu. Rev.
  • Substrate selection rules for the hairpin ribozyme determined by in vitro selection, mutation, and analysis of mismatched substrates. Genes Dev. (1993), 7(1), 130-8. xx x v Berzal-Herranz, Alfredo; Joseph, Simpson; Burke, John M.. In vitro selection of active hairpin ribozymes by sequential RNA-catalyzed cleavage and ligation reactions. Genes Dev. (1992), 6(1), 129-
  • Wait time does not include contact time during delivery.
  • AAAUAUUU A CAAGUACG 25 CGUACUUG CUGAUGAG GCCGUUAGGC CGAA AAAUAUUU 2214
  • AAACACAU C AGGUCAGG 42 CCUGACCU CUGAUGAG GCCGUUAGGC CGAA AUGUGUUU 2231
  • AAACCACU U CCGAUAAG 47 CUUAUCGG CUGAUGAG GCCGUUAGGC CGAA AGUGGUTJU 2236
  • AAAAGAUU A CCUGCAGC 251 GCUGCAGG CUGAUGAG GCCGUUAGGC CGAA AAUCUUUU 2440
  • AAAUCAUU C AUCCUUUU 531 AAAAGGAU CUGAUGAG GCCGUUAGGC CGAA AAUGAUUU 2720
  • AAAAUUUU C UAAAAUGU 545 ACAUUUUA CUGAUGAG GCCGUUAGGC CGAA AAAAUUUU 2734
  • NM_001285 Homo sapiens chloride channel, calcium activated, 1 (CLCAl) mRNA, 3311 bp

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Abstract

Nucleic acid molecules, including antisense and enzymatic nucleic acid molecules, such as hammerhead ribozymes, DNAzymes, and GeneBlocs, which modulate the expression of calcium activated chloride channel-1 (CLCA1).

Description

DESCRIPTION
METHOD AND REAGENT FOR THE INHIBITION OF CALCIUM ACTIVATED CHLORIDE CHANNEL-1 (CLCA-1)
Background Of The Invention
The present invention concerns compounds, compositions, and methods for the study, diagnosis, and treatment of conditions and diseases related to the expression of CLCA (Ch Channel Ca2+- Activated) genes.
The following is a brief description of the current understanding of CLCAs. The discussion is not meant to be complete and is provided only for understanding the invention that follows. The summary is not an admission that any of the work described below is prior art to the claimed invention.
CLCA proteins are emerging as a new class of channel proteins that mediate Ca2+-activated CI" conductance in a variety of tissues. Members of the CLCA family have been cloned, isolated, and partially characterized from human, bovine, and murine species. These proteins demonstrate a high degree of homology in their size, sequence, and predicted structure yet can vary considerably in tissue distribution. Bovine CLCAl (bCLCAl or CaCC) was the first reported CLCA homolog. The bCLCAl protein, which was isolated from and is exclusively detected in trachial epithelial cells, functions as a Ca2+-activated CI" channel (Ran > and Benos, 1992, J Biol. Chem., 267, 3618-3625; Cunningham et al, 1995, J. Biol. Chem., 270, 31016-31026). Another bovine homolog, bovine lung-endothelial cell adhesion molecule- 1 (Lu-ECAM-1), appears to have involvement in the preferential metastasis of melanoma cells to the lung. Lu-ECAM-1 shares 92% nucleotide identity to bCLCAl and is expressed in vascular endothelial cells (Elble et al, 1997, J. Biol Chem., 272, 27853-27861). It has been shown that Lu-ECAM-1, can mediate the binding of lung-metastatic mouse B16F10 melanoma cells to endothelial. cells (Zhu et al, 1992, J. Clin. Invest, 89, 1718-1724), however, due to sequence similarity to bCLCAl, the role of Lu-ECAM-1 as a chloride channel has been suggested (Elble et al, supra). The mouse homolog, mCLCAl, appears to have an expression pattern similar to the cystic fibrosis transmembrane conductance regulator (CFTR), with expression seen in various secretory epithelial cells,' squamous epithelia, and in some lymphocytes (Gruber et al, 1998, Histochem. Cell Biol, 110, 43-49). The three human CLCA homologs (hCLCAl, hCLCA2, and hCLCA3) thus far cloned, isolated, and partially characterized, all retain sequence homology, similar cDNA length, and are all located on the short arm of chromosome 1 (lp22- p31). Human CLCA proteins show a restricted pattern of expression in differing secretory tissues. Human CLCAl was the first reported calcium activated chloride channel in humans. The 31,902-bp hCLCAl gene is located on chromosome lp22- p31, contains 14 introns, and is preceded by a canonic promoter region that contains an LI transposable element. Expression of hCLCAl is predominant in intestinal basal crypt epithelia and goblet cells. A protein processing model has been proposed for hCLCAl in which the primary translation product (125-kDa) is cleaved to a 90- kDa and a group of 37- to 41 -kDa proteins, the latter apparently representing different glycosylation products of the same polypeptide (Gruber et al, 1998, Genomics, 54, 200-214); Transient expression of hCLCAl cDNA in HEK 293 cells is associated with an increase in whole-cell Ca2+-activated CI" conductance that is susceptible to inhibition with anion channel blocking compounds. Cell attached patch recordings of transfected cells in this study revealed single channels with a slope conductance of 13.4 pS (Gruber et al, supra).
The hCLCA2 homolog is processed in a similar manner as is hCLCAl, resulting in the formation of a heterodimer consisting of a 90-kDa amino terminal and an approximately 35 -kDa carboxy terminal subunit with anchorage to the plasma membrane via four or five transmembrane domains. Expression of hCLCA2 is somewhat less restricted than that of hCLCAl, being expressed from human lung, trachea, and breast tissue (Gruber et al, 1999, Am. J. Physiol, 276, C1261-C1270). Human CLCA2 is expressed in normal breast epithelium but not in breast tumors of different stages of progression, suggesting that hCLCA2 may act as a tumor suppressor in breast cancer (Gruber et al, 1999, Cancer Res., 59, 5488-5491). Human CLCA3 is a truncated, secreted member of the CLCA family which is expressed in numerous tissues including lung, trachea, spleen, thymus, and breast tissue. Unlike hCLCAl and hCLCA2 which are processed into heterodimers, hCLCA3 mRNA encodes a 37-kDa glycoprotein that corresponds to the N-terminal extracellular domain of its homologs. When hCLCA3 is expressed in HEK 293 or CHO cells, the 37-kDa glycoprotein is secreted (Gruber and Pauli, 1999, Biochem. Biophys. Acta, 1444, 418-423).
Holroyd et al, International PCT publication No. WO/9944620, describe a calcium-activated chloride channel that is induced by IL-9. Summary Of The Invention
The invention features novel nucleic acid-based techniques [e.g., enzymatic nucleic acid molecules (ribozymes), antisense nucleic acids, 2-5A antisense chimeras, triplex DNA, antisense nucleic acids containing RNA cleaving chemical groups] and methods for their use to modulate the expression of CLCA (CI" Channel
Ca2+-Activated) genes.
In a preferred embodiment, the invention features the use of one or more of the nucleic acid-based techniques independently or in combination to inhibit the expression of the genes encoding hCLCAl, hCLCA2, hCLCA3, and hCLCA4. Specifically, the invention features the use of nucleic acid-based techniques to specifically inhibit the expression of CLCAl (GenBank accession Nos. NM_001285, AF039400, AF039401, AF127036), CLCA2 (GenBank accession No. NM_006536), CLCA3 (GenBank accession No. NM_004921), and CLCA4 (GenBank accession No. NM_012128) genes. In yet another preferred embodiment, the invention features the inhibition of CLCAl gene using the nucleic acid-based techniques of the instant invention.
In another preferred embodiment, the invention features the use of an enzymatic nucleic acid molecule, preferably in the hammerhead, NCH (hiozyme), G- cleaver, amberzyme, zinzyme and/or DNAzyme motif, to inhibit the expression of CLCA genes.
By "inhibit" it is meant that the activity of CLCAl or level of RNAs or equivalent RNAs encoding one or more protein subunits of CLCAl is reduced below that observed in the-absence of the nucleic acid molecules of the invention. In one embodiment, inhibition with enzymatic nucleic acid molecules preferably is below that level observed in the presence of an enzymatically inactive or attenuated molecule that is able to bind to the same site on the target RNA, but is unable to cleave that RNA. In another embodiment, inhibition with antisense oligonucleotides is preferably below that level observed in the presence of, for example, an oligonucleotide with scrambled sequence or with mismatches. In another embodiment, inhibition of CLCAl genes with the nucleic acid molecule of the instant invention is greater than in the presence of the nucleic acid molecule than in its absence, or the presence of a control, irrelevant, or non-inhibitory oligonucleotide.
By "enzymatic nucleic acid molecule" it is meant a nucleic acid molecule which has complementarity in a substrate binding region to a specified gene target, and also has an enzymatic activity which is active to specifically cleave target RNA. That is, the enzymatic nucleic acid molecule is able to intermolecularly cleave RNA and thereby inactivate a target RNA molecule. These complementary regions allow sufficient hybridization of the enzymatic nucleic acid molecule to the target RNA and thus permit cleavage. One hundred percent complementarity is preferred, but complementarity as low as 50-75% may also be useful in this invention. The nucleic acids may be modified at the base, sugar, and/or phosphate groups. The term enzymatic nucleic acid is used interchangeably with phrases such as ribozymes, catalytic RNA, enzymatic RNA, catalytic DNA, aptazyme or aptamer-binding ribozyme, regulatable ribozyme, catalytic oligonucleotides, nucleozyme, DNAzyme, RNA enzyme, endoribonuclease, endonuclease, minizyme, leadzyme, oligozyme or DNA enzyme. All of these terminologies describe nucleic acid molecules with enzymatic activity. The specific enzymatic nucleic acid molecules described in the instant application are not meant to be limiting and those skilled in the art will recognize that all that is important in an enzymatic nucleic acid molecule of this invention is that it have a specific substrate binding site which is complementary to one or more of the target nucleic acid regions, and that it have nucleotide sequences within or surrounding that substrate binding site which impart a nucleic acid cleaving activity to the molecule (Cech et al, U.S. Patent No. 4,987,071; Cech et al, 1988, JAMA).
By "nucleic acid molecule" as used herein is meant a molecule having nucleotides. The nucleic acid can be single, double, or multiple stranded and may comprise modified or unmodified nucleotides or non-nucleotides or various mixtures and combinations thereof.
By "enzymatic portion" or "catalytic domain" is meant that portion/region of the enzymatic nucleic acid molecule essential for cleavage of a nucleic acid substrate (for example, see Figures 1-4).
By "substrate binding arm" or "substrate binding domain" is meant that portion/region of a ribozyme which is complementary to (i.e., able to base-pair with) a portion of its substrate. Generally, such complementarity is 100%, but can be less if desired. For example, as few as 10 bases out of 14 may be base-paired. Examples of such arms are shown generally in Figures 1-4. That is, these arms contain sequences within a ribozyme which are intended to bring ribozyme and target RNA together through complementary base-pairing interactions. The ribozyme of the invention may have binding arms that are contiguous or non-contiguous and may be of varying lengths. The length of the binding arm(s) are preferably greater than or equal to four nucleotides and of sufficient length to stably interact with the target RNA; specifically 12-100 nucleotides; more specifically 14-24 nucleotides long. If two binding arms are chosen, the design is such that the length of the binding arms are symmetrical (i.e., each of the binding arms is of the same length; e.g., five and five nucleotides, six and six nucleotides or seven and seven nucleotides long) or asymmetrical (i.e., the binding arms are of different length; e.g., six and three nucleotides; three and six nucleotides long; four and five nucleotides long; four and six nucleotides long; four and seven nucleotides long; and the like).
By "NCH" or "Inozyme" motif is meant, an enzymatic nucleic acid molecule comprising a motif as described in Ludwig et al, USSN No. 09/406,643, filed September 27, 1999, entitled "COMPOSITIONS HAVING RNA CLEANING ACTIVITY", and International PCT publication Nos. WO 98/58058 and WO 98/58057, all incorporated by reference herein in their entirety including the drawings.
By "G-cleaver" motif is meant, an enzymatic nucleic acid molecule comprising a motif as described in Eckstein et al, mtemational PCT publication No. WO 99/16871, incorporated by reference herein in its entirety including the drawings.
By "zinzyme" motif is meant, a class π enzymatic nucleic acid molecule comprising a motif as described in Beigelman et al, International PCT publication
No. WO 99/55857, incorporated by reference herein in its entirety including the drawings. Zinzymes represent a non-limiting example of an enzymatic nucleic acid molecule that does not require a ribonucleotide (2' -OH) group within its own nucleic
"acid sequence"fόr activity. ~
By "amberzyme" motif is meant, a class I enzymatic nucleic acid molecule comprising a motif as described in Beigelman et al, International PCT publication No. WO 99/55857, incorporated by reference herein in its entirety including the drawings. Amberzymes represent a non-limiting example of an enzymatic nucleic acid molecule that does not require a ribonucleotide (2'-OH) group within its own nucleic acid sequence for activity.
By 'DNAzyme' is meant, an enzymatic nucleic acid molecule that does not require the presence of a ribonucleotide (2' -OH) group within the DNAzyme molecule for its activity. In particular embodiments the enzymatic nucleic acid molecule may have an attached linker(s) or other attached or associated groups, moieties, or chains containing one or more nucleotides with 2'-OH groups. DNAzyme can be synthesized chemically or expressed endogenously in vivo, by means of a single stranded DNA vector or equivalent thereof.
By "sufficient length" is meant an oligonucleotide of greater than or equal to 3 nucleotides that is of a length great enough to provide the intended function under the expected condition. For example, for binding arms of enzymatic nucleic acid "sufficient length" means that the binding arm sequence is long enough to provide stable binding to a target site under the expected binding conditions. Preferably, the binding arms are not so long as to prevent useful turnover.
By "stably interact" is meant, interaction of the oligonucleotides with target nucleic acid (e.g., by forming hydrogen bonds with complementary nucleotides in the target under physiological conditions).
By "equivalent" RNA to CLCAl is meant to include those naturally occurring RNA molecules having homology (partial or complete) to CLCAl proteins or encoding for proteins with similar function as CLCAl in various organisms, including human, rodent, primate, rabbit, pig, protozoans, fungi, plants, and other microorganisms and parasites. The equivalent RNA sequence also includes in addition to the coding region, regions such as 5 '-untranslated region, 3 '-untranslated region, introns, intron-exon junction and the like.
By "homology" is meant the nucleotide sequence of two or more nucleic acid molecules is partially or completely identical.
By "antisense nucleic acid", it is meant a non-enzymatic nucleic acid molecule that binds to target RNA by means of RNA-RNA or RNA-DNA or RNA- PNA (protein nucleic acid; Egholm et al, 1993 Nature 365, 566) interactions and alters the activity of the target RNA (for a review, see Stein and Cheng, 1993 Science 261, 1004 and Woolf et al, US patent No. 5,849,902). Typically, antisense molecules will be complementary to a target sequence along a single contiguous sequence of the antisense molecule. However, in certain embodiments, an antisense molecule may bind to substrate such that the substrate molecule forms a loop, and/or an antisense molecule may bind such that the antisense molecule forms a loop. Thus, the antisense molecule may be complementary to two (or even more) noncontiguous substrate sequences or two (or even more) non-contiguous sequence portions of an antisense molecule may be complementary to a target sequence or both. For a review of current antisense strategies, see Schmajuk et al, 1999, J. Biol. Chem., 274, 21783-21789, Delihas et al, 1991, Nature, 15, 751-753, Stein et al, 1997, Antisense N A. Drug Dev., 7, 151, Crooke, 1998, Biotech. Genet. Eng. Rev., 15, 121-157, Crooke, 1997, Ad. Pharmacol, 40, 1-49. In addition, antisense DNA can be used to target RNA by means of DNA-RNA interactions, thereby activating RNase H, which digests the target RNA in the duplex. Antisense DNA can be synthesized chemically or expressed via the use of a single stranded DNA expression vector or equivalent thereof.
By "2-5A antisense chimera" it is meant, an antisense oligonucleotide containing a 5'-phosphorylated 2'-5'-linked adenylate residue. These chimeras bind to target RNA in a sequence-specific manner and activate a cellular 2-5A-dependent ribonuclease which, in turn, cleaves the target RNA (Torrence et al, 1993 Proc.
Natl. Acad. Sci. USA 90, 1300).
By "triplex DNA" it is meant an oligonucleotide that can bind to a double- stranded DNA in a sequence-specific manner to form a triple-strand helix. Formation of such triple helix structure has been shown to inhibit transcription of the targeted gene (Duval- Valentin et al, 1992 Proc. Natl Acad. Sci. USA 89, 504).
By "gene" it is meant a nucleic acid that encodes an RNA.
By "complementarity" is meant that a nucleic acid can form hydrogen bond(s) with another RNA sequence by either traditional Watson-Crick or other non- traditional types. In reference to the nucleic molecules of the present invention, the binding free energy for a nucleic acid molecule with its target or complementary sequence is sufficient to allow the relevant function of the nucleic acid to proceed, e.g., ribozyme cleavage, antisense or triple helix inhibition. Determination of binding free energies for nucleic acid molecules is well known in the art (see, e.g., Turner et al., 1987, CSHSymp. Quant. Biol. UL pp.123-133; Frier et al, 1986, Proc. Nat. Acad. Sci. USA 83:9373-9377; Turner et al, 1987, J. Am. Chem. Soc. 109:3783-3785). A percent complementarity indicates the percentage of contiguous residues in a nucleic acid molecule which can form hydrogen bonds (e.g., Watson- Crick base pairing) with a second nucleic acid sequence (e.g., 5, 6, 7, 8, 9, 10 out of 10 being 50%, 60%, 70%, 80%, 90%, and 100% complementary). "Perfectly complementary" means that all the contiguous residues of a nucleic acid sequence will hydrogen bond with the same number of contiguous residues in a second nucleic acid sequence.
At least seven basic varieties of naturally occurring enzymatic nucleic acids are known presently. Each can catalyze the hydrolysis of RNA phosphodiester bonds in trans (and thus can cleave other RNA molecules) under physiological conditions. Table I summarizes some of the characteristics of these ribozymes. In general, enzymatic nucleic acids act by first binding to a target RNA. Such binding occurs through the target binding portion of a enzymatic nucleic acid which is held in close proximity to an enzymatic portion of the molecule that acts to cleave the target RNA. Thus, the enzymatic nucleic acid first recognizes and then binds a target RNA through complementary base-pairing, and once bound to the correct site, acts enzymatically to cut the target RNA. Strategic cleavage of such a target RNA will destroy its ability to direct synthesis of an encoded protein. After an enzymatic nucleic acid has bound and cleaved its RNA target, it is released from that RNA to search for another target and can repeatedly bind and cleave new targets. Thus, a single ribozyme molecule is able to cleave many molecules of target RNA. In addition, the ribozyme is a highly specific inhibitor of gene expression, with the specificity of inhibition depending not only on the base-pairing mechanism of binding to the target RNA, but also on the mechanism of target RNA cleavage. Single mismatches, or base-substitutions, near the site of cleavage can completely eliminate catalytic activity of a ribozyme.
The enzymatic nucleic acid molecule that cleave the specified sites in CLCAl - specific RNAs represent a novel therapeutic approach to treat Chronic Obstructive
Pulmonary Diseases (COPDs), chronic bronchitis, asthma, cystic fibrosis, obstructive bowel syndrome, and other indications that may respond to the level of
CLCAl.
In one of the preferred embodiments of the inventions described herein, the enzymatic nucleic acid molecule is formed in a hammerhead or hairpin motif, but may also be formed in the motif of a hepatitis delta vims, group I intron, group π intron or RNase P RNA (in association with an RNA guide sequence), Neurospora VS RNA, DNAzymes, NCH cleaving motifs, or G-cleavers. Examples of such hammerhead motifs are described by Dreyfus, supra, Rossi et al, 1992, AIDS Research and Human Retroviruses 8, 183; Examples of hairpin motifs are described by Hampel et ah, EP0360257, Hampel and Tritz, 1989 Biochemistry 28, 4929, Feldstein et al, 1989, Gene 82, 53, Haseloff and Gerlach, 1989, Gene, 82, 43, Hampel et al, 1990 Nucleic Acids Res. 18, 299; Chowrira & McSwiggen, US. Patent No. 5,631,359. The hepatitis delta vims motif is described by Perrotta and Been, 1992 Biochemistry 31, 16. The RNase P motif is described by Guerrier- Takada et al, 1983 Cell 35, 849; Forster and Altman, 1990, Science 249, 783; Li and Altman, 1996, Nucleic Acids Res. 24, 835. Neurospora VS RNA ribozyme motif is described by Collins (Saville and Collins, 1990 Cell 61, 685-696; Saville and Collins, 1991 Proc. Natl. Acad. Sci. USA 88, 8826-8830; Collins and Olive, 1993 Biochemistry 32, 2795-2799; Guo and Collins, 1995, EMBO. J. 14, 363). Group II introns are described by Griffin et al, 1995, Chem. Biol. 2, 761; Michels and Pyle, 1995, Biochemistry 34, 2965; Pyle et al, International PCT Publication No. WO 96/22689. The Group I intron is described by Cech et al, U.S. Patent 4,987,071. DNAzymes are described by Usman et al, International PCT Publication No. WO 95/11304; Chartrand et ah, 1995, NAR 23, 4092; Breaker et al, 1995, Chem. Bio. 2, 655; Santoro et al, 1997, PNAS 94, 4262. NCH cleaving motifs are described in Ludwig & Sproat, International PCT Publication No. WO 98/58058; and G-cleavers are described in Kore et al., 1998, Nucleic Acids Research 26, 4116-4120 and Eckstein et al, International PCT Publication No. WO 99/16871. Additional motifs such as the Aptazyme (Breaker et al, WO 98/43993), Amberzyme (Class I motif; Figure 3; Beigelman et al, International PCT publication No. WO 99/55857) and Zinzyme (Beigelman et al, International PCT publication No. WO 99/55857), all these references are incorporated by reference herein in their totalities, including drawings and can also be used in the present invention. These specific motifs are not limiting in the invention, and those skilled in the art will recognize that all that is important in an enzymatic nucleic acid molecule of this invention is that it has a specific substrate binding site which is complementary to one or more of the target gene RNA regions, and that it have nucleotide sequences within or surrounding that substrate binding site which impart an RNA cleaving activity to the molecule (Cech et al, U.S. Patent No. 4,987,071).
In preferred embodiments of the present invention, a nucleic acid molecule, e.g., an antisense molecule, a triplex DNA, or a ribozyme, is 13 to 100 nucleotides in length, e.g., in specific embodiments 35, 36, 37, or 38 nucleotides in length (e.g., for particular ribozymes or. antisense).. _in particular embodiments, the nucleic acid molecule is 15-100, 17-100, 20-100, 21-100, 23-100, 25-100, 27-100, 30-100, 32- 100, 35-100, 40-100, 50-100, 60-100, 70-100, or 80-100 nucleotides in length. Instead of 100 nucleotides being the upper limit on the length ranges specified above, the upper limit of the length range can be, for example, 30, 40, 50, 60, 70, or 80 nucleotides. Thus, for any of the length ranges, the length range for particular embodiments has lower limit as specified, with an upper limit as specified which is greater than the lower limit. For example, in a particular embodiment, the length range can be 35-50 nucleotides in length. All such ranges are expressly included. Also in particular embodiments, a nucleic acid molecule can have a length which is any of the lengths specified above, for example, 21 nucleotides in length.
In a preferred embodiment, the invention provides a method for producing a class of nucleic acid-based gene inhibiting agents which exhibit a high degree of specificity for the RNA of a desired target. For example, the enzymatic nucleic acid molecule is preferably targeted to a highly conserved sequence region of target RNAs encoding CLCA proteins (for example, CLCAl, CLCA2, CLCA3 and/or CLCA4) such that specific treatment of a disease or condition can be provided with either one or several nucleic acid molecules of the invention. Such nucleic acid molecules can be delivered exogenously to specific tissue or cellular targets as required. Alternatively, the nucleic acid molecules (e.g., ribozymes and antisense) can be expressed from DNA and/or RNA vectors that are delivered to specific cells.
In a preferred embodiment, the invention features the use of nucleic acid-based inhibitors of the invention to specifically target genes that share homology with the CLCAl gene.
As used herein "cell" is used in its usual biological sense, and does not refer to an entire multicellular organism, e.g., specifically does not refer to a human. The cell may be present in a non-human multicellular organism, e.g., birds, plants and mammals such as cows, sheep, apes, monkeys, swine, dogs, and cats.
By "CLCA proteins" is meant, a protein or a mutant protein derivative thereof, comprising a calcium activated chloride channel protein.
By "highly conserved sequence region" is meant, a nucleotide sequence of one or more regions in a target gene does not vary significantly from one generation to the other or from one biological system to the other.
The nucleic acid-based inhibitors of CLCAl expression are useful for the prevention and/or treatment of diseases and conditions including Chronic Obstructive Pulmonary Disease (COPD), chronic bronchitis, asthma, cystic fibrosis, obstructive bowel syndrome, and any other diseases or conditions that are related to or will respond to the levels of CLCAl in a cell or tissue, alone or in combination with other therapies.
By "related" is meant that the reduction of CLCAl expression (specifically CLCAl gene) RNA levels and thus reduction in the level of the respective protein will relieve, to some extent, the symptoms of the disease or condition.
The nucleic acid-based inhibitors of the invention are added directly, or can be complexed with cationic lipids, packaged within liposomes, or otherwise delivered to target cells or tissues. The nucleic acid or nucleic acid complexes can be locally administered to relevant tissues ex vivo, or in vivo through injection, infusion pump or stent, with or without their incorporation in biopolymers. In preferred embodiments, the enzymatic nucleic acid inhibitors comprise sequences, which are complementary to the substrate sequences in Tables III to IX. Examples of such enzymatic nucleic acid molecules also are shown in Tables III to IX. Examples of such enzymatic nucleic acid molecules consist essentially of sequences defined in these Tables.
In yet another embodiment, the invention features antisense nucleic acid molecules and 2-5A chimera including sequences complementary to the substrate sequences shown in Tables III to IX. Such nucleic acid molecules can include sequences as shown for the binding arms of the enzymatic nucleic acid molecules in Tables III to VIII and sequences shown as GeneBloc™ sequences in Table IX. Similarly, triplex molecules can be provided targeted to the corresponding DNA target regions, and containing the DNA equivalent of a target sequence or a sequence complementary to the specified target (substrate) sequence. Typically, antisense molecules will be complementary to a target sequence along a single contiguous sequence of the antisense molecule. However, in certain embodiments, an antisense molecule may bind to substrate such that the substrate molecule forms a loop, and/or an antisense molecule may bind such that the antisense molecule forms a loop. Thus, the antisense molecule may be complementary to two (or even more) non- contiguous substrate sequences or two (or even more) non-contiguous sequence portions of an antisense molecule may be complementary to a target sequence or both.
By "consists essentially of is meant that the active nucleic acid molecule of the invention, for example, an enzymatic nucleic acid molecule, contains an enzymatic center or core equivalent to those in the examples, and binding arms able to bind RNA such that cleavage at the target site occurs. Other sequences can be present which do not interfere with such cleavage. Thus, a core region can, for example, include one or more loop, stem-loop structure, or linker which does not prevent enzymatic activity. Thus, the underlined regions in the sequences in Tables III, IV and VIII can be such a loop, stem-loop, nucleotide linker, and/or non- nucleotide linker and can be represented generally as sequence "X". For example, a core sequence for a hammerhead enzymatic nucleic acid can comprise a conserved sequence, such as 5'-CUGAUGAG-3' and 5'-CGAA-3' connected by "X", where X is 5'-GCCGUUAGGC-3' (SEQ ID NO 5450), or any other Stem π region known in the art. In another aspect of the invention, ribozymes or antisense molecules that interact with target RNA molecules and inhibit CLCAl (specifically CLCAl gene) activity are expressed from transcription units inserted into DNA or RNA vectors. The recombinant vectors are preferably DNA plasmids or viral vectors. Ribozyme or antisense expressing viral vectors could be constracted based on, but not limited to, adeno-associated virus, retrovirus, adenovirus, or alphavirus. Preferably, the recombinant vectors capable of expressing the ribozymes or antisense are delivered as described above, and persist in target cells. Alternatively, viral vectors may be used that provide for transient expression of ribozymes or antisense. Such vectors can be repeatedly administered as necessary. Once expressed, the ribozymes or antisense bind to the target RNA and inhibit its function or expression. Delivery of ribozyme or antisense expressing vectors can be systemic, such as by intravenous or intramuscular administration, by administration to target cells ex-planted from the patient followed by reintroduction into the patient, or by any other means that would allow for introduction into the desired target cell. Antisense DNA can be expressed endogenously via the use of a single stranded DNA intracellular expression vector.
By RNA is meant a molecule comprising at least one ribonucleotide residue. By "ribonucleotide" is meant a nucleotide with a hydroxyl group at the 2' position of a β-D-ribo-furanose moiety.
By "vectors" is meant any nucleic acid- and/or viral-based technique used to deliver a desired nucleic acid.
By "patient" is meant an organism, which is a donor or recipient of explanted cells or the cells themselves. "Patient" also refers to an organism to which the nucleic acid molecules of the invention can be administered. Preferably, a patient is a mammal or mammalian cells. More preferably, a patient is a human or human cells.
The nucleic acid molecules of the instant invention, individually, or in combination or in conjunction with other drugs, can be used to treat diseases or conditions discussed above. For example, to treat a disease or condition associated with the levels of CLCAl, the patient may be treated, or other appropriate cells may be treated, as is evident to those skilled in the art, individually or in combination with one or. more drugs under conditions suitable for the treatment.
In a further embodiment, the described molecules, such as antisense or ribozymes, can be used in combination with other known treatments to treat conditions or diseases discussed above. For example, the described molecules could be used in combination with one or more known therapeutic agents to treat Chronic Obstructive Pulmonary Diseases (COPDs), chronic bronchitis, asthma, cystic fibrosis, obstructive bowel syndrome, and/or other disease states or conditions which respond to the modulation of CLCAl expression.
In another preferred embodiment, the invention features nucleic acid-based inhibitors (e.g., enzymatic nucleic acid molecules (ribozymes), antisense nucleic acids, 2-5A antisense chimeras, triplex DNA, antisense nucleic acids containing RNA cleaving chemical groups) and methods for their use to down regulate or inhibit the expression of genes (e.g., CLCAl) capable of progression and/or maintenance of Chronic Obstructive Pulmonary Diseases (COPDs), chronic bronchitis, asthma, cystic fibrosis, obstructive bowel syndrome, and/or other disease states or conditions which respond to the modulation of CLCAl expression.
By "comprising" is meant including, but not limited to, whatever follows the word "comprising". Thus, use of the term "comprising" indicates that the listed elements are required or mandatory, but that other elements are optional and may or may not be present. By "consisting of is meant including, and limited to, whatever follows the phrase "consisting of. Thus, the phrase "consisting of indicates that the listed elements are required or mandatory, and that no other elements may be present. By "consisting essentially of is meant including any elements listed after the phrase, and limited to other elements that do not interfere with or contribute to the activity or action specified in the disclosure for the listed elements. Thus, the phrase "consisting essentially of indicates that the listed elements are required or mandatory, but that other elements are optional and may or may not be present depending upon whether or not they affect the activity or action of the listed elements.
The foregoing description of the various aspects and embodiments is provided with reference to the exemplary calcium activated chloride channel gene CLCAl, which is also referred to as CaCCl or ICACC-1. However, the various aspects and embodiments are also directed to other genes which express CLCAl or CaCCl-like proteins (for example hCLCA2, hCLCA3, hCLCA4, CaCC2, and CaCC3). Those additional genes can be analyzed for target sites using the methods described for CLCAl. Thus, the inhibition and the effects of such inhibition of the other genes can be performed as described herein.
Other features and advantages of the invention will be apparent from the following description of the preferred embodiments thereof, and from the claims. Description Of The Preferred Embodiments
First the drawings will be described briefly.
Drawings
Figure 1 shows examples of chemically stabilized ribozyme motifs. HH Rz, represents hammerhead ribozyme motif (Usman et al, 1996, Curr. Op. Struct. Bio., 1, 527); NCH Rz represents the NCH ribozyme motif (Ludwig & Sproat, Intemational PCT Publication No. WO 98/58058); G-CIeaver, represents G-cleaver ribozyme motif (Kore et al, 1998, Nucleic Acids Research 26, 4116-4120). N or n, represent independently a nucleotide which may be same or different and have complementarity to each other; ri, represents ribo-Inosine nucleotide; arrow indicates the site of cleavage within the target. Position 4 of the HH Rz and the NCH Rz is shown as having 2'-C-allyl modification, but those skilled in the art will recognize that this position can be modified with other modifications well known in the art, so long as such modifications do not significantly inhibit the activity of the ribozyme.
Figure 2 shows an example of the Amberzyme ribozyme motif that is chemically stabilized (see, for example, Beigelman et al, Intemational PCT publication No. WO 99/55857, incorporated by reference herein; also referred to as
Class I Motif). The Amberzyme motif is a class of enzymatic nucleic molecules that do not require the presence of a ribonucleotide (2'-OH) group for its activity.
Figure 3 shows an example of the Zinzyme A ribozyme motif that is chemically stabilized (Beigelman et al, International PCT publication No. WO
99/55857, incorporated by reference herein; also referred to as Class A or Class II
Motif). The Zinzyme motif is a class of enzymatic nucleic molecules that do not require the presence of a ribonucleotide (2' -OH) group for its activity.
Figure 4 shows an example of a DNAzyme motif described by Santoro et al., 1991, PNAS, 94, 4262.
Figures 5A and 5B are diagrammatic schemes representative of the process used for Target Discovery in the instant invention. The process for Target Discovery is described in Jarvis et al, Intemational PCT publication No. WO 98/50530, incorporated by reference herein in its entirety including the Figures.
Mechanism of action of Nucleic Acid Molecules of the Invention Antisense: Antisense molecules may be modified or unmodified RNA, DNA, or mixed polymer oligonucleotides which primarily function by specifically binding to matching sequences resulting in inhibition of peptide synthesis (Wu-Pong, Nov 1994, BioPharm, 20-33). The antisense oligonucleotide binds to target RNA by Watson Crick base-pairing and blocks gene expression by preventing ribosomal translation of the bound sequences either by steric blocking or by activating RNase H enzyme. Antisense molecules can also alter protein synthesis by interfering with RNA processing or transport from the nucleus into the cytoplasm (Mukhopadhyay & Roth, 1996, Crit. Rev. in Oncogenesis 7, 151-190).
In addition, binding of single stranded DNA to RNA may result in nuclease degradation of the heteroduplex (Wu-Pong, supra; Crooke, supra). To date, the only backbone modified DNA chemistry which will act as substrates for RNase H are phosphorothioates, phosphorodithioates, and borontrifluoridates. Recently it has been reported that 2'-arabino and 2'-fluoro arabino- containing oligos can also activate RNase H activity.
A number of antisense molecules have been described that utilize novel configurations of chemically modified nucleotides, secondary stracture, and/or RNase H substrate domains (Woolf et al, International PCT Publication No. WO 98/13526; Thompson et al, Intemational PCT Publication No. WO 99/54459; Hartmann et al, USSN 60/101,174 which was filed on September 21, 1998) all of these are incorporated by reference herein in their entirety.
In addition, antisense deoxyoligoribonucleotides can be used to target RNA by means of DNA-RNA interactions, thereby activating RNase H, which digests the target RNA in the duplex. Antisense DNA can be expressed endogenously in vivo via the use of a single stranded DNA intracellular expression vector or equivalents and variations thereof.
Triplex Forming Oligonucleotides (TFO : Single stranded DNA may be designed to bind to genomic DNA in a sequence specific manner. TFOs are comprised of pyrimidine-rich oligonucleotides which bind DNA helices through Hoogsteen Base-pairing (Wu-Pong, supra). The resulting triple helix composed of the DNA sense, DNA antisense, and TFO disrupts RNA synthesis by RNA polymerase. The TFO mechanism may result in gene expression or cell death since binding may be irreversible (Mukhopadhyay & Roth, supra).
2-5A Antisense Chimera: The 2-5A system is an interferon mediated mechanism for RNA degradation found in higher vertebrates (Mitra et al, 1996, Proc Nat Acad Sci USA 93, 6780-6785). Two types of enzymes, 2-5A synthetase and RNase L, are required for RNA cleavage. The 2-5A synthetases require double stranded RNA to form 2'-5' oligoadenylates (2-5 A). 2-5 A then acts as an allosteric effector for utilizing RNase L which has the ability to cleave single stranded RNA. The ability to form 2-5A structures with double stranded RNA makes this system particularly useful for inhibition of viral replication.
(2'-5') oligoadenylate structures may be covalently linked to antisense molecules to form chimeric oligonucleotides capable of RNA cleavage (Torrence, supra). These molecules putatively bind and activate a 2-5 A dependent RNase, the oligonucleotide/enzyme complex then binds to a target RNA molecule which can then be cleaved by the RNase enzyme.
Enzymatic Nucleic Acid: Seven basic varieties of naturally occurring enzymatic RNAs are presently known. In addition, several in vitro selection (evolution) strategies (Orgel, 1979, Proc. R. Soc. London, B 205, 435) have been used to evolve new nucleic acid catalysts capable of catalyzing cleavage and ligation of phosphodiester linkages (Joyce, 1989, Gene, 82, 83-87; Beaudry et al, 1992, Science 257, 635-641; Joyce, 1992, Scientific American 267, 90-97; Breaker et al, 1994, TIBTECH 12, 268; Bartel et al., 1993, Science 261:1411-1418; Szostak, 1993, IWS 17, 89-93; Kumar et al, 1995, FASEB J., 9, 1183; Breaker, 1996, Curr. Op. Biotech., 7, 442; Santoro et al, 1997, Proc. Natl Acad. Sci., 94, 4262; Tang et al, 1997, RNA 3, 914; Nakamaye & Eckstein, 1994, supra; Long & Uhlenbeck, 1994, supra; Ishizaka et al, 1995, supra; Vaish et al, 1997, Biochemistry 36, 6495; all of these are incoφorated by reference herein). Each can catalyze a series of reactions including the hydrolysis of phosphodiester bonds in trans (and thus can cleave other RNA molecules) under physiological conditions.
Nucleic acid molecules of this invention will block to some extent CLCAl protein expression and can be used to treat disease or diagnose disease associated with the levels of CLCAl.
The enzymatic nature of a ribozyme has significant advantages, such as the concentration of ribozyme necessary to affect a therapeutic treatment is lower. This advantage reflects the ability of the ribozyme to act enzymatically. Thus, a single ribozyme molecule is able to cleave many molecules of target RNA. In addition, the ribozyme is a highly specific inhibitor, with the specificity of inhibition depending not only on the base-pairing mechanism of binding to the target RNA, but also on the mechanism of target RNA cleavage. Single mismatches, or base-substitutions, near the site of cleavage can be chosen to completely eliminate catalytic activity of a ribozyme.
Nucleic acid molecules having an endonuclease enzymatic activity are able to repeatedly cleave other separate RNA molecules in a nucleotide base sequence- specific manner. Such enzymatic nucleic acid molecules can be targeted to virtually any RNA transcript, and achieve efficient cleavage in vitro (Zaug et al, 324, Nature 429 1986 ; Uhlenbeck, 1987 Nature 328, 596; Kim et al, 84 Proc. Natl Acad. Sci. USA 8788, 1987; Dreyfus, 1988, Einstein Quart. J. Bio. Med., 6, 92; Haseloff and Gerlach, 334 Nature 585, 1988; Cech, 260 JAMA 3030, 1988; and Jefferies et al, 17 Nucleic Acids Research 1371, 1989; Santoro et al, 1997 supra).
Because of their sequence specificity, trans-cleaving ribozymes show promise as therapeutic agents for human disease (Usman and McSwiggen, 1995 Ann. Rep. Med. Chem. 30, 285-294; Christoffersen and Marr, 1995 J. Med. Chem. 38, 2023- 2037). Ribozymes can be designed to cleave specific RNA targets within the background of cellular RNA. Such a cleavage event renders the RNA nonfunctional and abrogates protein expression from that RNA. In this manner, synthesis of a protein associated with a disease state can be selectively inhibited (Warashina et al, 1999, Chemistry and Biology, 6, 237-250).
The nucleic acid molecules of the instant invention are also referred to as GeneBloc reagents, which are essentially nucleic acid molecules (e.g.; ribozymes, antisense) capable of down-regulating gene expression.
GeneBlocs are modified oligonucleotides including ribozymes and modified antisense oligonucleotides that bind— to and target specific mRNA molecules. Because GeneBlocs can be designed to target any specific mRNA, their potential applications are quite broad. Traditional antisense approaches have often relied heavily on the use of phosphorothioate modifications to enhance stability in biological samples, leading to a myriad of specificity problems stemming from nonspecific protein binding and general cytotoxicity (Stein, 1995, Nature Medicine, 1, 1119). In contrast, GeneBlocs contain a number of modifications that confer nuclease resistance while making minimal use of phosphorothioate linkages, which reduces toxicity, increases binding affinity and minimizes non-specific effects compared with traditional antisense oligonucleotides. Similar reagents have recently been utilized successfully in various cell culture systems (Vassar, et al, 1999, Science, 286, 735) and in vivo (Jarvis et al., manuscript in preparation). In addition, novel cationic lipids can be utilized to enhance cellular uptake in the presence of serum. Since ribozymes and antisense oligonucleotides regulate gene expression at the RNA level, the ability to maintain a steady-state dose of GeneBloc over several days was important for target protein and phenotypic analysis. The advances in resistance to nuclease degradation and prolonged activity in vitro have supported the use of GeneBlocs in target validation applications.
Target sites
Targets for useful ribozymes and antisense nucleic acids can be determined as disclosed in Draper et al, WO 93/23569; Sullivan et al, WO 93/23057; Thompson et al, WO 94/02595; Draper et al, WO 95/04818; McSwiggen et al, US Patent No. 5,525,468. All of these publications are hereby incorporated by reference herein in their totality. Other examples include the following PCT applications, which concern inactivation of expression of disease-related genes: WO 95/23225, WO 95/13380, WO 94/02595, all of which are incorporated by reference herein. Rather than repeat the guidance provided in those documents here, specific examples of such methods are provided herein, not limiting to those in the art. Ribozymes and antisense to such targets are designed as described in those applications and synthesized to be tested in vitro and in vivo, as also described. The sequences of human CLCAl RNAs were screened for optimal enzymatic nucleic acid and antisense target sites using a computer-folding algorithm. Antisense, hammerhead, DNAzyme, NCH, amberzyme, zinzyme, or G-Cleaver ribozyme binding/cleavage sites were identified. These sites are shown in Tables III to IX (all sequences are 5' to 3' in the tables; the underlined region can be any base-paired sequence, the actual sequence is not relevant here). The nucleotide base position is noted in the Tables as that site to be cleaved by the designated type of enzymatic nucleic acid molecule. While human sequences can be screened and enzymatic nucleic acid molecule and/or antisense thereafter designed, as discussed in Stinchcomb et al, WO 95/23225, mouse targeted ribozymes may be useful to test efficacy of action of the enzymatic nucleic acid molecule and/or antisense prior to testing in humans.
Antisense, hammerhead, DNAzyme, NCH, amberzyme, zinzyme or G-Cleaver ribozyme binding/cleavage sites were identified. The nucleic acid molecules are individually analyzed by computer folding (Jaeger et al, 1989 Proc. Natl Acad. Sci. USA, 86, 7706) to assess whether the sequences fold into the appropriate secondary stracture. Those nucleic acid molecules with unfavorable intramolecular interactions such as between the binding arms and the catalytic core are eliminated from consideration. Varying binding arm lengths can be chosen to optimize activity. Antisense, hammerhead, DNAzyme, NCH, amberzyme, zinzyme or G-Cleaver ribozyme binding/cleavage sites were identified and were designed to anneal to various sites in the RNA target. The binding arms are complementary to the target site sequences described above. The nucleic acid molecules were chemically synthesized. The method of synthesis used follows the procedure for normal DNA/RNA synthesis as described below and in Usman et al, 1987 J. Am. Chem. Soc, 109, 7845; Scaringe et al, 1990 Nucleic Acids Res., 18, 5433; Wincott et al, 1995 Nucleic Acids Res. 23, 2677-2684; and Carafhers et al., 1992, Methods in Enzymology 211,3-19.
Synthesis of Nucleic acid Molecules
Synthesis of nucleic acids greater than 100 nucleotides in length is difficult using automated methods, and the therapeutic cost of such molecules is prohibitive. In this invention, small nucleic acid motifs ("small refers to nucleic acid motifs no more than 100 nucleotides in length, preferably no more than 80 nucleotides in length, and most preferably no more than 50 nucleotides in length; e.g., antisense oligonucleotides, hammerhead or the NCH ribozymes) are preferably used for exogenous delivery. The simple structure of these molecules increases the ability of the nucleic acid to invade targeted regions of RNA structure. Exemplary molecules of the instant invention are chemically synthesized, and others can similarly be synthesized.
Oligonucleotides (e.g.; antisense GeneBlocs) are synthesized using protocols known in the art as described in Carathers et al, 1992, Methods in Enzymology 211, 3-19, Thompson et al, Intemational PCT Publication No. WO 99/54459, Wincott et al, 1995, Nucleic Acids Res. 23, 2677-2684, Wincott et al, 1997, Methods Mol. Bio., 74, 59, Brennan et al, 1998, Biotechnol Bioeng., 61, 33-45, and Brennan, US patent No. 6,001,311. All of these references are incorporated herein by reference. The synthesis of oligonucleotides makes use of common nucleic acid protecting and coupling groups, such as dimethoxytrityl at the 5'-end, and phosphoramidites at the 3'-end. In a non-limiting example, small scale syntheses are conducted on a 394 Applied Biosystems, Inc. synthesizer using a 0.2 μmol scale protocol with a 2.5 min coupling step for 2'-O-methylated nucleotides and a 45 sec coupling step for 2'- deoxy nucleotides. Table II outlines the amounts and the contact times of the reagents used in the synthesis cycle. Alternatively, syntheses at the 0.2 μmol scale can be performed on a 96-well plate synthesizer, such as the instrument produced by Protogene (Palo Alto, CA) with minimal modification to the cycle. A 33-fold excess (60 μL of 0.11 M •= 6.6 μmol) of 2'-O-methyl phosphoramidite and a 105-fold excess of S-ethyl tetrazole (60 μL of 0.25 M = 15 μmol) can be used in each coupling cycle of 2'-O-methyl residues relative to polymer-bound 5 '-hydroxyl. A 22-fold excess (40 μL of 0.11 M = 4.4^ μmol) of deoxy phosphoramidite and a 70- fold excess of S-ethyl tetrazole (40 μL of 0.25 M = 10 μmol) can be used in each coupling cycle of deoxy residues relative to polymer-bound 5 '-hydroxyl. Average coupling yields on the 394 Applied Biosystems, Inc. synthesizer, determined by colorimetric quantitation of the trityl fractions, are typically 97.5-99%. Other oligonucleotide synthesis reagents for the 394 Applied Biosystems, Inc. synthesizer include; detritylation solution is 3% TCA in methylene chloride (ABI); capping is performed with 16% N-methyl imidazole in THF (ABI) and 10% acetic anhydride/10% 2,6-lutidine in THF (ABI); and oxidation solution is 16.9 mM 12, 49 mM pyridine, 9% water in THF (PERSEPTIVE™). Burdick & Jackson Synthesis Grade acetonitrile is used directly from the reagent bottle. S-Ethyltetrazole solution (0.25 M in acetonitrile) is made up from the solid obtained from American International Chemical, Inc. Alternately, for the introduction of phosphorothioate linkages, Beaucage reagent (3H-l,2-Benzodithiol-3-one 1,1 -dioxide, 0.05 M in acetonitrile) is used.
Deprotection of the antisense oligonucleotides is performed as follows: the polymer-bound trityl-on oligoribonucleotide is transferred to a 4 mL glass screw top vial and suspended in a solution of 40% aq. methylamine (1 mL) at 65 °C for 10 min. After cooling to -20 °C, the supernatant is removed from the polymer support. The support is washed three times with 1.0 mL of EtOH:MeCΝ:H2O/3:l:l, vortexed and the supernatant is then added to the first supernatant. The combined supernatants, containing the oligoribonucleotide, are dried to a white powder.
The method of synthesis used for normal RNA including certain enzymatic nucleic acid molecules follows the procedure as described in Usman et al, 1987, J. Am. Chem. Soc, 109, 7845; Scaringe et al, 1990, Nucleic Acids Res., 18, 5433; Wincott et al, 1995, Nucleic Acids Res. 23, 2677-2684 and Wincott et al, 1997, Methods Mol. Bio., 74, 59, and makes use of common nucleic acid protecting and coupling groups, such as dimethoxytrityl at the 5'-end, and phosphoramidites at the 3'-end. In a non-limiting example, small scale syntheses are conducted on a 394 Applied Biosystems, Inc. synthesizer using a 0.2 μmol scale protocol with a 7.5 min coupling step for alkylsilyl protected nucleotides and a 2.5 min coupling step for 2'- O-methylated nucleotides. Table II outlines the amounts and the contact times of the reagents used in the synthesis cycle. Alternatively, syntheses at the 0.2 μmol scale can be done on a 96-well plate synthesizer, such as the instrument produced by Protogene (Palo Alto, CA) with minimal modification to the cycle. A 33-fold excess (60 μL of 0.11 M = 6.6 μmol) of 2'-O-methyl phosphoramidite and a 75-fold excess of S-ethyl tetrazole (60 μL of 0.25 M = 15 μmol) can be used in each coupling cycle of 2'-O-methyl residues relative to polymer-bound 5'-hydroxyl. A 66-fold excess (120 μL of 0.11 M = 13.2 μmol) of alkylsilyl (ribo) protected phosphoramidite and a 150-fold excess of S-ethyl tetrazole (120 μL of 0.25 M = 30 μmol) can be used in each coupling cycle of rib residues relative to polymer-bound 5 '-hydroxyl. Average coupling yields on the 394 Applied Biosystems, Inc. synthesizer, determined by colorimetric quantitation of the trityl fractions, are typically 97.5- 99%. Other oligonucleotide synthesis reagents for the 394 Applied Biosystems, Inc. synthesizer include; detritylation solution is 3% TCA in methylene chloride (ABI); capping is performed with 16% N-methyl imidazole in THF (ABI) and 10% acetic anhydride/10% 2,6-lutidine in THF (ABI); oxidation solution is 16.9 mM Ϊ2, 49 mM pyridine, 9% water in THF (PERSEPTIVE™). Burdick & Jackson Synthesis Grade acetonitrile is used directly from the reagent bottle. S-Ethyltetrazole solution (0.25 M in acetonitrile) is made up from the solid obtained from American Intemational Chemical, Inc. Alternately, for the introduction of phosphorothioate linkages, Beaucage reagent (3H-l,2-Benzodifhiol-3-one l,l-dioxide0.05 M in acetonitrile) is used.
Deprotection of the RΝA is performed using either a two-pot or one-pot protocol. For the two-pot protocol, the polymer-bound trityl-on oligoribonucleotide is transferred to a 4 mL glass screw top vial and suspended in a solution of 40% aq. methylamine (1 mL) at 65 °C for 10 min. After cooling to -20 °C, the supernatant is removed from the polymer support. The support is washed three times with 1.0 mL of EtOH:MeCΝ:H2O/3:l:l, vortexed and the supernatant is then added to the first supernatant. The combined supernatants, containing the oligoribonucleotide, are dried to a white powder. The base deprotected oligoribonucleotide is resuspended in anhydrous TEA HF/NMP solution (300 μL of a solution of 1.5 mL N- methylpyrrolidinone, 750 μL TEA and 1 mL TEA-3HF to provide a 1.4 M HF concentration) and heated to 65 °C. After 1.5 h, the oligomer is quenched with 1.5 M NΗ4HCO3.
Alternatively, for the one-pot protocol, the polymer-bound trityl-on oligoribonucleotide is transferred to a 4 mL glass screw top vial and suspended in a solution of 33% ethanolic methylamine/DMSO: 1/1 (0.8 mL) at 65 °C for 15 min.
The vial is brought to r.t. TEA«3HF (0.1 mL) is added and the vial is heated at 65 °C for 15 min. The sample is cooled at -20 °C and then quenched with 1.5 M NH4HCO3. For purification of the trityl-on oligomers, the quenched NH4HCO3 solution is loaded onto a C-18 containing cartridge that had been prewashed with acetonitrile followed by 50 mM TEAA. After washing the loaded cartridge with water, the RNA is detritylated with 0.5% TFA for 13 min. The cartridge is then washed again with water, salt exchanged with 1 M NaCl and washed with water again. The oligonucleotide is then eluted with 30% acetonitrile.
Inactive hammerhead ribozymes or binding attenuated control (BAC) oligonucleotides) are synthesized by substituting a U for G5 and a U for A14
(numbering from Hertel, K. J., et al, 1992, Nucleic Acids Res_., 20, 3252). Similarly, one or more nucleotide substitutions can be introduced in other enzymatic nucleic acid molecules to inactivate the molecule and such molecules can serve as a negative control.
The average stepwise coupling yields are typically >98% (Wincott et al, 1995
Nucleic Acids Res. 23, 2677-2684). Those of ordinary skill in the art will recognize that the scale of synthesis can be adapted to be larger or smaller than the examples described above including but not limited to 96-well format, all that is important is the ratio of chemicals used in the reaction.
Alternatively, the nucleic acid molecules of the present invention can be synthesized separately and joined together post- synthetically, for example by ligation (Moore et al, 1992, Science 256, 9923; Draper et al, Intemational PCT publication No. WO 93/23569; Shabarova et al, 1991, Nucleic Acids Research 19, 4247; Bellon et al, 1997, Nucleosides & Nucleotides, 16, 951; Bellon et al, 1997, Bioconjugate Chem. 8, 204).
The nucleic acid molecules of the present invention are modified extensively to enhance stability by modification with nuclease resistant groups, for example, 2'- amino, 2'-C-allyl, 2'-flouro, 2'-O-mefhyl, 2'-H (for a review see Usman and Cedergren, 1992, TIBS 17, 34; Usman et al, 1994, Nucleic Acids Symp. Ser. 31, 163). Ribozymes are purified by gel electrophoresis using general methods or are purified by high pressure liquid chromatography (HPLC; See Wincott et al, supra, the totality of which is hereby incoφorated herein by reference) and are resuspended in water.
The sequences of the ribozymes and antisense constructs that are chemically synthesized, useful in this study, are shown in Tables III to IX. Those in the art will recognize that these sequences are representative only of many more such sequences where the enzymatic portion of the ribozyme (all but the binding arms) is altered to affect activity. The ribozyme and antisense constract sequences listed in Tables III to IX may be formed of ribonucleotides or other nucleotides or non- nucleotides. Such ribozymes with enzymatic activity are equivalent to the ribozymes described specifically in the Tables. Optimizing Activity of the nucleic acid molecule of the invention.
Chemically synthesizing nucleic acid molecules with modifications (base, sugar and/or phosphate) that prevent their degradation by seram ribonucleases may increase their potency (see e.g., Eckstein et al, International Publication No. WO 92/07065; Perrault et al, 1990 Nature 344, 565; Pieken et al, 1991, Science 253, 314; Usman and Cedergren, 1992, Trends in Biochem. Sci. 17, 334; Usman et al, International Publication No. WO 93/15187; Rossi et al, Intemational Publication No. WO 91/03162; Sproat, US Patent No. 5,334,711; and Burgin et al, supra; all of these describe various chemical modifications that can be made to the base, phosphate and/or sugar moieties of the nucleic acid molecules described herein. All these references are incoφorated by reference herein. Modifications which enhance their efficacy in cells, and removal of bases from nucleic acid molecules to shorten oligonucleotide synthesis times and reduce chemical requirements are desired.
There are several examples in the art describing sugar, base and phosphate modifications that can be introduced into nucleic acid molecules with significant enhancement in their nuclease stability and efficacy. For example, oligonucleotides are modified to enhance stability and/or enhance biological activity by modification with nuclease resistant groups, for example, 2'-amino, 2'-C-allyl, 2'-flouro, -O- methyl, 2'-H, nucleotide base modifications (for a review see Usman and Cedergren, 1992, TIBS. 17, 34; Usman et al, 1994, Nucleic Acids Symp. Ser. 31, 163; Burgin et al, 1996, Biochemistry , 35, 14090). Sugar modifications of nucleic acid molecules have been extensively described in the art (see Eckstein et al, International Publication PCT No. WO 92/07065; Perrault et al. Nature, 1990, 344, 565-568; Pieken et al. Science, 1991, 253, 314-317; Usman and Cedergren, Trends in Biochem. Sci. , 1992, 17, 334-339; Usman et al. International Publication PCT No. WO 93/15187; Sproat, US Patent No. 5,334,711 and Beigelman et al, 1995, J Biol Chem., 270, 25702; Beigelman et al, Intemational PCT publication No. WO 97/26270; Beigelman et al, US Patent No. 5,716,824; Usman et al, US patent No. 5,627,053; Woolf et al, Intemational PCT Publication No. WO 98/13526; Thompson et al, USSN 60/082,404 which was filed on April 20, 1998; Kaφeisky et al, 1998, Tetrahedron Lett., 39, 1131; Earnshaw and Gait, 1998, Biopolymers (Nucleic acid Sciences), 48, 39-55; Verma and Eckstein, 1998, Annu. Rev. Biochem., 67, 99-134; and Burlina et al, 1997, Bioorg. Med. Chem., 5, 1999-2010; all of the references are hereby incoφorated by reference herein in their totalities). Such publications describe general methods and strategies to determine the location of incoφoration of sugar, base and or phosphate modifications and the like into ribozymes without inhibiting catalysis. In view of such teachings, similar modifications can be used as described herein to modify the nucleic acid molecules of the instant invention.
While chemical modification of oligonucleotide intemucleotide linkages with phosphorothioate, phosphorothioate, and or 5'-methylphosphonate linkages improves stability, too many of these modifications may cause some toxicity. Therefore when designing nucleic acid molecules the amount of these intemucleotide linkages should be minimized. The reduction in the concentration of these linkages should lower toxicity resulting in increased efficacy and higher specificity of these molecules.
Nucleic acid molecules having chemical modifications which maintain or enhance activity are provided. Such nucleic acid is also generally more resistant to nucleases than unmodified nucleic acid. Thus, in a cell and/or in vivo the activity may not be significantly lowered. Therapeutic nucleic acid molecules delivered exogenously must optimally be stable within cells until translation of the target RNA has been inhibited long enough to reduce the levels of the undesirable protein. This period of time varies between hours to days depending upon the disease state. Clearly, nucleic acid molecules must be resistant to nucleases in order to function as effective intracellular therapeutic agents. Improvements in the chemical synthesis of RNA and DNA (Wincott et al, 1995 Nucleic Acids Res. 23, 2677; Carafhers et al, 1992, Methods in Enzymology 211,3-19 (incoφorated by reference herein) have expanded the ability to modify nucleic acid molecules by introducing nucleotide modifications to enhance their nuclease stability as described above.
Use of these the nucleic acid-based molecules of the invention will lead to better treatment of the disease progression by affording the possibility of combination therapies (e.g., multiple antisense or enzymatic nucleic acid molecules targeted to different genes, nucleic acid molecules coupled with known small molecule inhibitors, or intermittent treatment with combinations of molecules (including different motifs) and/or other chemical or biological molecules). The treatment of patients with nucleic acid molecules may also include combinations of different types of nucleic acid molecules. Therapeutic nucleic acid molecules (e.g., enzymatic nucleic acid molecules and antisense nucleic acid molecules) delivered exogenously must optimally be stable within cells until translation of the target RNA has been inhibited long enough to reduce the levels of the undesirable protein. This period of time varies between hours to days depending upon the disease state. Clearly, these nucleic acid molecules must be resistant to nucleases in order to function as effective intracellular therapeutic agents. Improvements in the chemical synthesis of nucleic acid molecules described in the instant invention and in the art have expanded the ability to modify nucleic acid molecules by introducing nucleotide modifications to enhance their nuclease stability as described above.
By "enhanced enzymatic activity" is meant to include activity measured in cells and/or in vivo where the activity is a reflection of both catalytic activity and ribozyme stability. In this invention, the product of these properties is increased or not significantly (less than 10- fold) decreased in vivo compared to an all RNA ribozyme or all DNA enzyme.
In yet another preferred embodiment, nucleic acid catalysts having chemical modifications which maintain or enhance enzymatic activity are provided. Such nucleic acid is also generally more resistant to nucleases than unmodified nucleic acid. Thus, in a cell and/or in vivo the activity may not be significantly lowered. As exemplified herein such ribozymes are useful in a cell and/or in vivo even if activity over all is reduced 10 fold (Burgin et al, 1996, Biochemistry, 35, 14090). Such ribozymes herein are said to "maintain" the enzymatic activity of an all RNA ribozyme.
In another aspect the nucleic acid molecules comprise a 5' and/or a 3'- cap structure.
By "cap stracture" is meant chemical modifications, which have been incoφorated at either terminus of the oligonucleotide (see, for example, Wincott et al, WO 97/26270, incoφorated by reference herein). These terminal modifications protect the nucleic acid molecule from exonuclease degradation, and may help in delivery and/or localization within a cell. The cap may be present at the 5 '-terminus (5 '-cap) or at the 3 '-terminus (3 '-cap) or may be present on both termini. In non- limiting examples the 5 '-cap is selected from the group comprising inverted abasic residue (moiety), 4',5'-methylene nucleotide; l-(beta-D-erythrofuranosyl) nucleotide, 4'-thio nucleotide, , carbocyclic nucleotide; 1,5-anhydrohexitol nucleotide; L- nucleotides; alpha-nucleotides; modified base nucleotide; phosphorodithioate linkage; t/zreo-pentofuranosyl nucleotide; acyclic 3',4'-seco nucleotide; acyclic 3,4- dihydroxybutyl nucleotide; acyclic 3,5-dihydroxypentyl nucleotide, 3'-3 '-inverted nucleotide moiety; 3 '-3 '-inverted abasic moiety; 3'-2'-inverted nucleotide moiety; 3'- 2'-inverted abasic moiety; 1,4-butanediol phosphate; 3'-phosphoramidate; hexylphosphate; aminohexyl phosphate; 3'-phosphate; 3'-phosphorothioate; phosphorodithioate; or bridging or non-bridging methylphosphonate moiety (for more details see Wincott et al, Intemational PCT publication No. WO 97/26270, incoφorated by reference herein).
In yet another preferred embodiment, the 3 '-cap is selected from a group comprising, 4',5'-mefhylene nucleotide; l-(beta-D-erythrofuranosyl) nucleotide; 4'- thio nucleotide, carbocyclic nucleotide; 5'-amino-alkyl phosphate; l,3-diamino-2- propyl phosphate, 3-aminopropyl phosphate; 6-aminohexyl phosphate; 1,2- aminododecyl phosphate; hydroxypropyl phosphate; 1,5-anhydrohexitol nucleotide;
L-nucleotide; alpha-nucleotide; modified base nucleotide; phosphorodithioate; t/zreo-pentofuranosyl nucleotide; acyclic 3',4'-seco nucleotide; 3,4-dihydroxybutyl nucleotide; 3,5-dihydroxypentyl nucleotide, 5'-5'-inverted nucleotide moiety; 5'-5'- inverted abasic moiety; 5'-phosphoramidate; 5'-phosphorothioate; 1,4-butanediol phosphate; 5'-amino; bridging and/or non-bridging 5'-phosphoramidate, phosphorothioate and/or phosphorodithioate, bridging or non bridging methylphosphonate and 5'-mercapto moieties (for more details, see Beaucage and
Iyer, 1993, Tetrahedron 49, 1925; incoφorated by reference herein).
By the term "non-nucleotide" is meant any group or compound which can be incoφorated into a nucleic acid chain in the place of one or more nucleotide units, including either sugar and/or phosphate substitutions, and allows the remaining bases to exhibit their enzymatic activity. The group or compound is abasic in that it does not contain a commonly recognized nucleotide base, such as adenosine, guanine, cytosine, uracil or thymine.
An "alkyl" group refers to a saturated aliphatic hydrocarbon, including straight-chain, branched-chain, and cyclic alkyl groups. Preferably, the alkyl group has 1 to 12 carbons. More preferably it is a lower alkyl of from 1 to 7 carbons, more preferably 1 to 4 carbons. The alkyl group may be substituted or unsubstituted. When substituted the substituted group(s) is preferably, hydroxyl, cyano, alkoxy, =O, =S, NO2 or N(CH3)2, amino, or SH. The term also includes alkenyl groups which are unsaturated hydrocarbon groups containing at least one carbon-carbon double bond, including straight-chain, branched-chain, and cyclic groups.
Preferably, the alkenyl group has 1 to 12 carbons. More preferably it is a lower alkenyl of from 1 to 7 carbons, more preferably 1 to 4 carbons. The alkenyl group may be substituted or unsubstituted. When substituted the substituted group(s) is preferably, hydroxyl, cyano, alkoxy, =O, =S, NO2, halogen, N(CH3)2, amino, or
SH. The term "alkyl" also includes alkynyl groups which have an unsaturated hydrocarbon group containing at least one carbon-carbon triple bond, including straight-chain, branched-chain, and cyclic groups. Preferably, the alkynyl group has 1 to 12 carbons. More preferably it is a lower alkynyl of from 1 to 7 carbons, more preferably 1 to 4 carbons. The alkynyl group may be substituted or unsubstituted. When substituted the substituted group(s) is preferably, hydroxyl, cyano, alkoxy, =O, =S, NO2 or N(CH3)2, amino or SH.
Such alkyl groups may also include aryl, alkylaryl, carbocyclic aryl, heterocyclic aryl, amide and ester groups. An "aryl" group refers to an aromatic group which has at least one ring having a conjugated π electron system and includes carbocyclic aryl, heterocyclic aryl and biaryl groups, all of which may be optionally substituted. The preferred substituent(s) of aryl groups are halogen, trihalomethyl, hydroxyl, SH, OH, cyano, alkoxy, alkyl, alkenyl, alkynyl, and amino groups. An "alkylaryl" group refers to an alkyl group (as described above) covalently joined to an aryl group (as described above). Carbocyclic aryl groups are groups wherein the ring atoms on the aromatic ring are all carbon atoms. The carbon atoms are optionally substituted. Heterocyclic aryl groups are groups having from 1 to 3 heteroatoms as ring atoms in the aromatic ring and the remainder of the ring atoms are carbon atoms. Suitable heteroatoms include oxygen, sulfur, and nitrogen, and include furanyl, thienyl, pyridyl, pyrrolyl, N-lower alkyl pyrrolo, pyrimidyl, pyrazinyl, imidazolyl and the like, all optionally substituted. An "amide" refers to an -C(O)-NH-R,. where R is either alkyl, aryl, alkylaryl or hydrogen. An "ester" refers to an -C(O)-OR', where R is either alkyl, aryl, alkylaryl or hydrogen.
By "nucleotide" as used herein is as recognized in the art to include natural bases (standard), and modified bases well known in the art. Such bases are generally located at the 1' position of a nucleotide sugar moiety. Nucleotides generally comprise a base, sugar and a phosphate group. The nucleotides can be unmodified or modified at the sugar, phosphate and/or base moiety, (also referred to interchangeably as nucleotide analogs, modified nucleotides, non-natural nucleotides, non-standard nucleotides and other; see for example, Usman and McSwiggen, supra; Eckstein et al, International PCT Publication No. WO 92/07065 ; Usman et al, Intemational PCT Publication No. WO 93/15187; Uhlmann & Peyman, 1990, Chemical Reviews, 90, 4, 544-579, all are hereby incoφorated by reference herein). There are several examples of modified nucleic acid bases known in the art as summarized by Limbach et al, 1994, Nucleic Acids Res. 22, 2183. Some of the non-limiting examples of base modifications that can be introduced into nucleic acid molecules include, inosine, purine, pyridin-4-one, pyridin-2-one, phenyl, pseudouracil, 2, 4, 6-trimethoxy benzene, 3 -methyl uracil, dihydrouridine, naphthyl, aminophenyl, 5-alkylcytidines (e.g., 5-methylcytidine), 5-alkyluridines (e.g., ribothymidine), 5-halouridine (e.g., 5-bromouridine) or 6-azapyrimidines or 6- alkylpyrimidines (e.g. 6-methyluridine), propyne, and others (Burgin et al, 1996, Biochemistry, 35, 14090; Uhlman & Peyman, supra). By "modified bases" in this aspect is meant nucleotide bases other than adenine, guanine, cytosine and uracil at 1' position or their equivalents; such bases may be used at any position, for example, within the catalytic core of an enzymatic nucleic acid molecule and/or in the substrate-binding regions of the nucleic acid molecule.
In a preferred embodiment, the invention features modified ribozymes with phosphate backbone modifications comprising one or more phosphorothioate, phosphorodithioate, methylphosphonate, moφholino, amidate carbamate, carboxymethyl, acetamidate, polyamide, sulfonate, sulfonamide, sulfamate, formacetal, thioformacetal, and/or alkylsilyl, substitutions. For a review of oligonucleotide backbone modifications see Hunziker and Leumann, 1995, Nucleic Acid Analogues: Synthesis and Properties, in Modern Synthetic Methods, VCH, 331-417, and Mesmaeker et al, 1994, Novel Backbone Replacements for Oligonucleotides, in Carbohydrate Modifications in Antisense Research, ACS, 24- 39. These references are hereby incoφorated by reference herein.
By "abasic" is meant sugar moieties lacking a base or having other chemical groups in place of a base at the 1' position, (for more details, see Wincott et al, Intemational PCT publication No. WO 97/26270).
By "unmodified nucleoside" is meant one of the bases adenine, cytosine, guanine, thymine, uracil joined to the 1' carbon of β-D-ribo-furanose.
By "modified nucleoside" is meant any nucleotide base which contains a modification in the chemical structure of an unmodified nucleotide base, sugar and/or phosphate.
In connection with 2 '-modified nucleotides as described for the present invention, by "amino" is meant 2'-NH2 or 2'-O- NH , which may be modified or unmodified. Such modified groups are described, for example, in Eckstein et al,
U.S. Patent 5,672,695 and Matulic-Adamic et al, WΟ 98/28317, respectively, which are both incoφorated by reference herein in their entireties. Various modifications to nucleic acid (e.g., antisense and ribozyme) structure can be made to enhance the utility of these molecules. Such modifications will enhance shelf-life, half-life in vitro, stability, and ease of introduction of such oligonucleotides to the target site, e.g., to enhance penetration of cellular membranes, and confer the ability to recognize and bind to targeted cells.
Use of these molecules will lead to better treatment of the disease progression by affording the possibility of combination therapies (e.g., multiple ribozymes targeted to different genes, ribozymes coupled with known small molecule inhibitors, or intermittent treatment with combinations of ribozymes (including different ribozyme motifs) and/or other chemical or biological molecules). The treatment of patients with nucleic acid molecules may also include combinations of different types of nucleic acid molecules. Therapies may be devised which include a mixture of ribozymes (including different ribozyme motifs), antisense and/or 2-5A chimera molecules to one or more targets to alleviate symptoms of a disease.
Administration of Nucleic Acid Molecules
Methods for the delivery of nucleic acid molecules are described in Akhtar et al, 1992, Trends Cell Bio., 2, 139; and Delivery Strategies for Antisense Oligonucleotide Therapeutics, ed. Akhtar, 1995 which are both incoφorated herein by reference. Sullivan et al, PCT WO 94/02595, further describes the general methods for delivery of enzymatic RNA molecules. These protocols may be utilized for the delivery of virtually any nucleic acid molecule. Nucleic acid molecules may be administered to cells by a variety of methods known to those familiar to the art, including, but not restricted to, encapsulation in liposomes, by iontophoresis, or by incoφoration into other vehicle's" such as hydrogels, cyclodextrins, biodegradable nanocapsules, and bioadhesive microspheres. For some- indications, nucleic acid molecules may be directly delivered ex vivo to cells or tissues with or without the aforementioned vehicles. Alternatively, the nucleic acid/vehicle combination is locally delivered by direct injection or by use of a catheter, infusion pump or stent. Other routes of delivery include, but are not limited to, intravascular, intramuscular, subcutaneous or joint injection, aerosol inhalation, oral (tablet or pill form), topical, systemic, ocular, intraperitoneal and/or infrathecal delivery. More detailed descriptions of nucleic acid delivery and administration are provided in Sullivan et al, supra, Draper et al, PCT WO93/23569, Beigelman et al, PCT WO99/05094, and Klimuk et al, PCT WO99/04819 all of which have been incoφorated by reference herein. In addition, the nucleic acid molecules of the instant invention, used to treat pulmonary diseases and disorders, may be administered directly to the lungs via pulmonary delivery. The pulmonary delivery of oligonucleotides is described by Bennett et al, Intemational PCT publication Nos. WO/9960166 and WO/9960010; Danahay et al, 1999, Pharm. Res., 16(10), 1542-1549; Metzger and Nyce, 1999, J. Allergy Clin. Immunol, 104(2, Pt. 1), 260-266; Nicklin et al, 1998, Pharm. Res., 15(4), 583-591; Ilium and Watts, Intemational PCT publication No. WO/9735562; and Nyce, 1991, Expert Opm. Invest. Drugs, 6(9), 1149-1156.
The molecules of the instant invention can be used as pharmaceutical agents. Pharmaceutical agents prevent, inhibit the occurrence, or treat (alleviate a symptom to some extent, preferably all of the symptoms) of a disease state in a patient.
The negatively charged polynucleotides of the invention can be administered (e.g., RNA, DNA or protein) and introduced into a patient by any standard means, with or without stabilizers, buffers, and the like, to form a pharmaceutical composition. When it is desired to use a liposome delivery mechanism, standard protocols for formation of liposomes can be followed. The compositions of the present invention may also be formulated and used as tablets, capsules or elixirs for oral administration; suppositories for rectal administration; sterile solutions; suspensions for injectable administration; and other compositions known in the art.
The present invention also includes pharmaceutically acceptable formulations of the compounds described. These formulations include salts of the above compounds, e.g., acid addition salts, including salts of hydrochloric, hydrobromic, acetic acid, and benzene sulfonic acid.
A pharmacological composition or formulation refers to a composition or formulation in a form suitable for administration, e.g., systemic administration, into a cell or patient, preferably a human. Suitable forms, in part, depend upon the use or the route of entry, for example oral, transdermal, or by injection. Such forms should not prevent the composition or formulation from reaching a target cell (i.e., a cell to which the negatively charged polymer is desired to be delivered to). For example, pharmacological compositions injected into the blood stream should be soluble.
Other factors are known in the art, and include considerations such as toxicity and forms which prevent the composition or formulation from exerting its effect.By
"systemic administration" is meant in vivo systemic absoφtion or accumulation of drags in the blood stream followed by distribution throughout the entire body. Administration routes that lead to systemic absoφtion include, without limitations: intravenous, subcutaneous, intraperitoneal, inhalation, oral, intrapulmonary and intramuscular. Each of these administration routes exposes the desired negatively charged polymers, e.g., nucleic acids, to an accessible diseased tissue. The rate of entry of a drug into the circulation has been shown to be a function of molecular weight or size. The use of a liposome or other drug carrier comprising the compounds of the instant invention can potentially localize the drag, for example, in certain tissue types, such as the tissues of the reticular endothelial system (RES). A liposome formulation that can facilitate the association of drug with the surface of cells, such as, lymphocytes and macrophages is also useful. This approach may provide enhanced delivery of the drug to target cells by taking advantage of the specificity of macrophage and lymphocyte immune recognition of abnormal cells, such as cancer cells.
By pharmaceutically acceptable formulation is meant, a composition or formulation that allows for the effective distribution of the nucleic acid molecules of the instant invention in the physical location most suitable for their desired activity. Non-limiting examples of agents suitable for formulation with the nucleic acid molecules of the instant invention include: P-glycoprotein inhibitors (such as Pluronic P85) which can enhance entry of drags into the CNS (Jolliet-Riant and Tillement, 1999, Fundam. Clin. Pharmacol, 13, 16-26); biodegradable polymers, such as poly (DL-lactide-coglycolide) microspheres for sustained release delivery after intracerebral implantation (Emerich, DF et al, 1999, Cell Transplant, 8, 47-58) Alkermes, Inc. Cambridge, MA; and loaded nanoparticles, such as those made of polybutylcyanoacrylate, which can deliver drags across the blood brain barrier and can alter neuronal uptake mechanisms (Prog Neuropsychopharmacol Biol Psychiatry, 23, 941-949, 1999). Other non-limiting examples of delivery strategies for the nucleic acid molecules of the instant invention include material described in Boado et al, 1998, J. Pharm. Sci., 87, 1308-1315; Tyler et al, 1999, FEBS Lett, 421, 280-284; Pardridge et al, 1995, PNAS USA., 92, 5592-5596; Boado, 1995, Adv. Drug Delivery Rev., 15, 73-107; Aldrian-Herrada et al, 1998, Nucleic Acids Res., 26, 4910-4916; and Tyler et al, 1999, PNAS USA., 96, 7053-7058.
The invention also features the use of the composition comprising surface- modified liposomes containing poly (ethylene glycol) lipids (PEG-modified, or long- circulating liposomes or stealth liposomes). These formulations offer a method for increasing the accumulation of drags in target tissues. This class of drag carriers resists opsonization and elimination by the mononuclear phagocytic system (MPS or RES), thereby enabling longer blood circulation times and enhanced tissue exposure for the encapsulated drag (Lasic et al. Chem. Rev. 1995, 95, 2601-2627; Ishiwata et al, Chem. Pharm. Bull 1995, 43, 1005-1011). All incoφorated by reference herein. Such liposomes have been shown to accumulate selectively in tumors, presumably by extravasation and capture in the neovascularized target tissues (Lasic et al, Science 1995, 267, 1275-1276; Oku et al, 1995, Biochim. Biophys. Acta, 1238, 86- 90). All incoφorated by reference herein. The long-circulating liposomes enhance the pharmacokinetics and pharmacodynamics of DNA and RNA, particularly compared to conventional cationic liposomes which are known to accumulate in tissues of the MPS (Liu et al, J. Biol. Chem. 1995, 42, 24864-24870; Choi et al, International PCT Publication No. WO 96/10391; Ansell et al, International PCT Publication No. WO 96/10390; Holland et al, Intemational PCT Publication No. WO 96/10392; all of which are incoφorated by reference herein). Long-circulating liposomes are also likely to protect drags from nuclease degradation to a greater extent compared to cationic liposomes, based on their ability to avoid accumulation in metabolically aggressive MPS tissues such as the liver and spleen.
In addition, the invention features the use of methods to deliver the nucleic acid molecules of the instant invention to hematopoietic cells, including monocytes and lymphocytes. These methods are described in detail by Hartmann et al, 1998, J. Phamacol Exp. Ther., 285(2), 920-928; Kronenwett et al, 1998, Blood, 91(3), 852- 862; Filion and Phillips, 1997, Biochim. Biophys. Acta., 1329(2), 345-356; Ma and Wei, 1996, Leuk. Res., 20(11/12), 925-930; and Bongartz et al, 1994, Nucleic Acids Research, 22(22), 4681-8. Such methods, as described above, include the use of free oligonucleotide, cationic lipid formulations, liposome formulations including pH sensitive liposomes and immunoliposomes, and bioconjugates including oligonucleotides conjugated to fusogenic peptides, for the transfection of hematopoietic cells with oligonucleotides.
The present invention also includes compositions prepared for storage or administration which include a pharmaceutically effective amount of the desired compounds in a pharmaceutically acceptable carrier or diluent. Acceptable carriers or diluents for therapeutic use are well known in the pharmaceutical art, and are described, for example, in Remington 's Pharmaceutical Sciences, Mack Publishing Co. (A.R. Gennaro edit. 1985) hereby incoφorated by reference herein. For example, preservatives, stabilizers, dyes and flavoring agents may be provided. These include sodium benzoate, sorbic acid and esters of j9-hydroxybenzoic acid. In addition, antioxidants and suspending agents may be used.
A pharmaceutically effective dose is that dose required to prevent, inhibit the occurrence, or treat (alleviate a symptom to some extent, preferably all of the symptoms) of a disease state. The pharmaceutically effective dose depends on the type of disease, the composition used, the route of administration, the type of mammal being treated, the physical characteristics of the specific mammal under consideration, concurrent medication, and other factors which those skilled in the medical arts will recognize. Generally, an amount between 0.1 mg/kg and 100 mg/kg body weight/day of active ingredients is administered dependent upon potency of the negatively charged polymer.
The nucleic acid molecules of the present invention may also be administered to a patient in combination with other therapeutic compounds to increase the overall therapeutic effect. The use of multiple compounds to treat an indication may increase the beneficial effects while reducing the presence of side effects. Oxygen therapy, bronchodilators, corticosteroids, antibacterials, vaccinations, acetylcysteine, mucokinetic agents, and DNase (Pulmozyme) are non-limiting examples of compounds and/or methods that can be combined with or used in conjunction with the nucleic acid molecules (e.g. ribozymes and antisense molecules) of the instant invention. Those skilled in the art will recognize that other drug compounds and therapies can be similarly and readily combined with the nucleic acid molecules of the instant invention (e.g. ribozymes and antisense molecules) and are, therefore, within the scope of the instant invention.
Alternatively, certain of the nucleic acid molecules of the instant invention can be expressed within cells from eukaryotic promoters (e.g., Izant and Weintraub, 1985, Science, 229, 345; McGarry and Lindquist, 1986, Proc. Natl Acad. Sci., USA 83, 399; Scanlon et al, 1991, Proc. Natl. Acad. Sci. USA, 88, 10591-5; Kashani- Sabet et al, 1992, Antisense Res. Dev., 2, 3-15; Dropulic et al, 1992, J. Virol, 66, 1432-41; Weerasinghe et al, 1991, J. Virol, 65, 5531-4; Ojwang et al, 1992, Proc. Natl. Acad. Sci. USA, 89, 10802-6; Chen et al, 1992, Nucleic Acids Res., 20, 4581-9; Sarver et al, 1990 Science, 247, 1222-1225; Thompson et al, 1995, Nucleic Acids Res., 23, 2259; Good et al, 1991, Gene Therapy, 4, 45; all of the references are hereby incoφorated in their totality by reference herein). Those skilled in the art realize that any nucleic acid can be expressed in eukaryotic cells from the appropriate DNA/RNA vector. The activity of such nucleic acids can be augmented by their release from the primary transcript by a ribozyme (Draper et al, PCT WO 93/23569, and Sullivan et al, PCT WO 94/02595; Ohkawa et al, 1992, Nucleic Acids Symp. Ser., 27, 15-6; Taira et α/., 1991, Nucleic Acids Res., 19, 5125- 30; Ventura et al, 1993, Nucleic Acids Res., 21, 3249-55; Chowrira et al, 1994, J. Biol. Chem., 269, 25856; all of these references are hereby incoφorated in their totalities by reference herein). In another aspect of the invention, RNA molecules of the present invention are preferably expressed from transcription units (see, for example, Couture et al, 1996, TIG., 12, 510) inserted into DNA or RNA vectors. The recombinant vectors are preferably DNA plasmids or viral vectors. Ribozyme expressing viral vectors could be constructed based on, but not limited to, adeno-associated virus, retrovirus, adenovirus, or alphaviras. Preferably, the recombinant vectors capable of expressing the nucleic acid molecules are delivered as described above, and persist in target cells. Alternatively, viral vectors may be used that provide for transient expression of nucleic acid molecules. Such vectors might be repeatedly administered as necessary. Once expressed, the nucleic acid molecule binds to the target mRNA. Delivery of nucleic acid molecule expressing vectors could be systemic, such as by intravenous or intra-muscular administration, by administration to target cells ex- planted from the patient followed by reintroduction into the patient, or by any other means that would allow for introduction into the desired target cell (for a review, see Couture et al, 1996, TIG., 12, 510).
In one aspect, the invention features an expression vector comprising a nucleic acid sequence encoding at least one of the nucleic acid molecules disclosed in the instant invention. The nucleic acid sequence encoding the nucleic acid molecule of the instant invention is operably linked in a manner which allows expression of that nucleic acid molecule.
In another aspect, the invention features an expression vector comprising: a) a transcription initiation region (e.g., eukaryotic pol I, II or in initiation region); b) a transcription termination region (e.g., eukaryotic pol I, π or HI termination region); c) a nucleic acid sequence encoding at least one of the nucleic acid catalyst of the instant invention; and wherein said sequence is operably linked to said initiation region and said termination region, in a manner which allows expression and/or delivery of said nucleic acid molecule. The vector may optionally include an open reading frame (ORF) for a protein operably linked on the 5' side or the 3'-side of the sequence encoding the nucleic acid catalyst of the invention; and/or an intron (intervening sequences).
Transcription of the nucleic acid molecule sequences are driven from a promoter for eukaryotic RNA polymerase I (pol I), RNA polymerase II (pol TL), or RNA polymerase HI (pol HI). Transcripts from pol H or pol HI promoters will be expressed at high levels in all cells; the levels of a given pol H promoter in a given cell type will depend on the nature of the gene regulatory sequences (enhancers, silencers, etc.) present nearby. Prokaryotic RNA polymerase promoters are also used, providing that the prokaryotic RNA polymerase enzyme is expressed in the appropriate cells (Elroy-Stein and Moss, 1990, Proc. Natl. Acad. Sci. U S A, 87, 6743-7; Gao and Huang 1993, Nucleic Acids Res.., 21, 2867-72; Lieber et al., 1993, Methods Enzymol, 217, 47-66; Zhou et al, 1990, Mol. Cell. Biol, 10, 4529- 37). All of these references are incoφorated by reference herein.
Several investigators have demonstrated that nucleic acid molecules, such as ribozymes expressed from such promoters can function in mammalian cells (e.g. Kashani-Sabet et al, 1992, Antisense Res. Dev., 2, 3-15; Ojwang et al, 1992, Proc. Nat Acad. Sci. USA, 89, 10802-6; Chen etal, 1992, Nucleic Acids Res., 20, 4581-9; Yu et al, 1993, Proc. Natl. Acad. Sci. USA, 90, 6340-4; L'Huillier et al, 1992, EMBO J., 11, 4411-8; Lisziewicz et al, 1993, Proc. Natl. Acad. Sci. U. S. A, 90, 8000-4; Thompson et. al, 1995, Nucleic Acids Res., 23, 2259; and Sullenger & Cech, 1993, Science, 262, 1566). More specifically, transcription units such as the ones derived from genes encoding U6 small nuclear (snRNA), transfer RNA (tRNA) and adenovirus VA RNA are useful in generating high concentrations of desired RNA molecules such as ribozymes in cells (Thompson et al, supra; Couture and Stinchcomb, 1996, supra; Noonberg et al, 1994, Nucleic Acid Res., 22, 2830; Noonberg et al, US Patent No. 5,624,803; Good et al, 1997, Gene Ther., 4, 45; and Beigelman et al, Intemational PCT Publication No. WO 96/18736; all of these publications are incoφorated by reference herein. The above ribozyme transcription units can be incoφorated into a variety of vectors for introduction into mammalian cells, including but not restricted to, plasmid DNA vectors, viral DNA vectors (such as adenovirus or adeno-associated virus vectors), or viral RNA vectors (such as retroviral or alphaviras vectors) (for a review, see Couture and Stinchcomb, 1996, supra).
In yet another aspect, the invention features an expression vector comprising a nucleic acid sequence encoding at least one of the nucleic acid molecules of the invention, in a manner which allows expression of that nucleic acid molecule. The expression vector comprises in one embodiment; a) a transcription initiation region; b) a transcription termination region; c) a nucleic acid sequence encoding at least one said nucleic acid molecule; and wherein said sequence is operably linked to said initiation region and said termination region, in a manner which allows expression and/or delivery of said nucleic acid molecule.
hi another preferred embodiment, the expression vector comprises: a) a transcription initiation region; b) a transcription termination region; c) an open reading frame; d) a nucleic acid sequence encoding at least one said nucleic acid molecule, wherein said sequence is operably linked to the 3 '-end of said open reading frame; and wherein said sequence is operably linked to said initiation region, said open reading frame and said termination region, in a manner which allows expression and/or delivery of said nucleic acid molecule.
In yet another embodiment the expression vector comprises: a) a transcription initiation region; b) a transcription termination region; c) an intron; d) a nucleic acid sequence encoding at least one said nucleic acid molecule; and wherein said sequence is operably linked to said initiation region, said intron and said termination region, in a manner which allows expression and or delivery of said nucleic acid molecule.
In another embodiment, the expression vector comprises: a) a transcription initiation region; b) a transcription termination region; c) an intron; d) an open reading frame; e) a nucleic acid sequence encoding at least one said nucleic acid molecule, wherein said sequence is operably linked to the 3'-end of said open reading frame; and wherein said sequence is operably linked to said initiation region, said intron, said open reading frame and said termination region, in a manner which allows expression and/or delivery of said nucleic acid molecule.
Examples.
The following are non-limiting examples showing the selection, isolation, synthesis and activity of nucleic acids of the instant invention.
The following examples demonstrate the selection and design of Antisense, hammerhead, DNAzyme, NCH, Amberzyme, Zinzyme, or G-Cleaver ribozyme molecules and binding/cleavage sites within CLCAl RNA.
Example 1: Reporter System
Applicant used a target discovery and target validation approach to finding genes that are involved in chronic mucous hypersecretion. In order to discover genes playing a role in the expression of mucins, a readily assayable reporter system was devised. The reporter system consists of a plasmid construct, termed pMUC5AC- EGFP, bearing a gene coding for Green Fluorescent Protein (GFP). The promoter region of the GFP gene is replaced by a portion of the Mucin 5AC promoter sufficient to direct efficient transcription of the GFP gene. The plasmid also contains the neomycin drug resistance gene. Example 2: Host Cell Line for Target Discovery
The cell line selected as host for these studies, NCI-H292 (ATCC CRL-1848), is derived from a human lung mucoepidermoid carcinoma. The cells retain mucoepidermoid characteristics in culture and endogenously express mucin 5AC and mucin 2. The ρMUC5AC-EGFP plasmid was transfected into NCI-H292 using a cationic lipid formulation. Following transfection, the cells were subjected to limiting dilution cloning under selection by 600 μg/mL Geneticin. Cells retaining the pMUC5AC-EGFP plasmid survive the Geneticin treatment and form colonies derived from single surviving cells.The resulting clonal cell lines were screened by flow cytometry for the capacity to upregulate GFP production directed by the Mucin 5AC promoter. Treating the cells with sterilized M9 bacterial medium in which Pseudomonas aeruginosa had been cultured (Pseudomonas conditioned medium, PCM) induced the mucin promoter. The PCM is supplemented with phorbol myristate acetate (PMA).
A clonal cell line highly responsive to mucin promoter induction, designated
H292/MUC5AC/EGFP Clone8 (H292 Clone 8) was selected as the reporter line for subsequent studies. The process for Target Discovery is described in Jarvis et al, International PCT publication No. WO 98/50530, incoφorated by reference herein in its entirety including the Figures.
Example 3: Ribozyme Library Constraction
A ribozyme library was constracted with oligonucletides containing ribozymes with two randomized regions comprising six-nucleotide binding "arms" (Stem I and Stem HI of a ribozyme-substrate complex). Oligo sequence 5' and 3' of the ribozyme contains restriction endonuclease cleavage sites for cloning. The 3' trailing sequence forms a stem-loop for priming DNA polymerase extension to form a double stranded molecule. The double-stranded ribozyme library was cloned into the U6+27 transcription unit located in the 5' LTR region of a retroviral vector containing the human nerve growth factor receptor (hNGFr) reporter gene. Positioning the U6+27/ribozyme transcription unit in the 5' LTR results in a duplication of the transcription unit when the vector integrates into the host cell genome. As a result, the ribozyme is transcribed by RNA polymerase HI from U6+27 and by RNA polymerase H activity directed by the 5' LTR. The ribozyme library was packaged into retroviral particles that were used to infect and transduce H292 Clone 8 cells. Assay of the hNGFr reporter indicated that 50% to 60% of Clone 8 cells incoφorated the ribozyme construct. Figure 5A and 5B describe the generalized scheme used in the ribozyme library constraction and target discovery. By "randomized region" is meant a region of completely random sequence and/or partially random sequence. By completely random sequence is meant a sequence wherein theoretically there is equal representation of A, T, G and C nucleotides or modified derivatives thereof, at each position in the sequence. By partially random sequence is meant a sequence wherein there is an unequal representation of A, T, G and C nucleotides or modified derivatives thereof, at each position in the sequence. A partially random sequence can therefore have one or more positions of complete randomness and one or more positions with defined nucleotides.
Example 4: Enriching for Non-responders to Mucin Induction
Sorting of ribozyme library-containing cells was performed to enrich for cells that produce less GFP after treatment with PCM and PMA. Lower GFP production may be due to ribozyme action upon genes involved in the activation of the mucin promoter. Alternatively, ribozymes may directly target the mucin/GFP transcript resulting in reduced GFP expression.
Cells were seeded at a density of 1 x 10 per 150 cm style cell culture flasks. After 72 hours the standard cell culture medium was replaced with medium without fetal bovine serum. After 24 hours of seram deprivation the cells were treated with serum-containing medium supplemented with PCM (to 40%) and PMA (to 50 nM) to induced GFP production via the mucin promoter. After 20 to 22 hours, cells were monitored for GFP level on a FACStar Plus cell sorter.
Sorting was performed if 90% of ribozyme library cells from an unsorted control sample were induced to produce GFP above background levels. Two cell fractions were collected in each round of sorting.
In the initial sort the Ml gate collected cells in luminescence channels 1 to 4.5; those cells with the lowest GFP signal (5% of the induced population). The M2 sort gate collected cells in luminescence channels 4.5 to 20; cells with low GFP signal (10% of the induced population). The Ml and M2 fractions together represented the 15% of the induced population responding least to the GFP induction treatment. In order to assure that the diversity of the ribozyme library was represented 2.3 X 106 cells were collected in the Ml fraction and 4.6 x 106 cells were collected in the M2 fraction. The Ml and M2 fractions wee cultured separately and representative portions of each were cryopreserved after each round of sorting. When treated with PCM and PMA prior to a second round of sorting, cells from both the Ml and M2 fractions responded as before with >90% of the cells producing elevated levels of GFP. The same sorting criteria and sort gates were used in the second round. As in the first round of sorting the Ml sort gate collected 5% of the treated cells (those with little or no GFP) and the M2 gate collected 10% of the cells. Two more rounds of sorting were performed using the same sorting criteria.
Prior to the third round of sorting the Ml fraction showed a three-fold enrichment of GFP negative cells. Prior to the fourth round of sorting both the Ml and M2 fractions were significantly enriched in cells unresponsive to the GFP induction treatment.
Following the third round of sorting the Ml fraction was selected to generate a database of ribozymes present in the sorted cells.
Example 5: Recovery of Ribozyme Sequence from Sorted Cells
Genomic DNA was obtained from sorted ribozyme library cells by standard methods. Nested polymerase chain reaction (PCR) primers (Sequence ID Nos. 5468 and 5469) that hybridized to the retroviral vector 5' and 3' of the ribozyme were used to recover and amplify the ribozyme sequences from the Clone 8 library cell DNA. The PCR product was ligated into a bacterial cloning vector. Two methods were developed to use the recovered ribozyme library, in plasmid form, to generate a database of ribozyme binding arm sequences. In the first approach the library was cloned into E. coli. DNA was prepared by plasmid isolation from bacterial colonies or by direct colony. PCR and _ribozyme..arm sequence.was determined. Over 450 sequences have been obtained by this method. A second method used the ribozyme library to transfect H292 Clone 8 cells. Clonal lines of stably transfected cells were established and induced with PCM and PMA. Those lines which failed to respond to GFP induction were probed by PCR for single ribozyme integration events. Over 300 sequences were obtained in this manner. The unique ribozyme sequences obtained by both methods were added to a Target Sequence Tag (TST) database.
Example 6: Bioinformatics
After sequencing 760 recovered ribozymes 171 unique sequences were found. Of the unique sequences, 91 have been recovered once and 80 have been found multiple times. Most of the repeated sequences have been found 2 to 11 times. One sequence has been recovered 145 times. The diversity of the sequences obtained indicates that the sorted cells are a promising source of information leading to target discovery.
Ribozyme binding arm sequences were compared to public and private gene data banks. Gene matches were compiled according to perfect and imperfect matches. Potential gene targets were categorized by the number of different ribozyme sequences matching each gene. Multiple ribozyme matches have been found for 180 genes. Genes with more than one perfect ribozyme match were given close attention. A total of 34 genes have been verified to date to have multiple perfect ribozyme matches. Of those at least 17 have protein products of known function.
Two perfect ribozyme matches were found for human calcium activated chloride channel-1 (hCLCAl). Each ribozyme matches at two sites in the hCLCAl gene. A third sorted library ribozyme sequence "hits" hCLCAl but has a single nucleotide mismatch.
Example 7: Selection of hCLCAl for Validation
The selection of hCLCAl as a candidate for target validation was based on bioinformatics and on emerging data in murine models of mucous hypersecretion in the trachea and lung. Two ribozymes (Seq. ID Nos. 2332 and 2273) recovered from cells that no longer respond to mucin promoter/GFP induction match perfectly to hCLCAl. A third has a single mismatch. Evidence from two murine models indicates a correlation between mucous hypersecretion in the lung and strong upregulation of gob-5 (GenBank ABO17156), a murine homologue of hCLCAl.
Example 8: Validation of hCLCAl
To validate hCLCAl as a regulator of MUC5AC expression, GeneBIoc reagents were designed (Table IX) to the hCLCAl cDNA sequence (GenBank AF039400). GeneBIoc reagents are complexed with a cationic lipid formulation prior to administration to H292/MUC5AC/GFP Clone 8 cells. Concentrations of the GeneBIoc reagents administered range from 30 nM to 120 nM at cationic lipid concentrations of 4-6 μg/mL. Cells are treated with GeneBIoc reagents for 72 to 96 hours. Before the termination of GeneBIoc treatment, PCM (to 40 %) and PMA (to 50 nM) are added to induce the MUC5AC promoter. After twenty hours of induction the cells are harvested and assayed for phenotypic and molecular parameters. Reduced GFP expression in GeneBIoc treated cells (measured by flow cytometry) is taken as evidence for validation of hCLCAl. Knockdown of hCLCAl RNA in GeneBIoc treated cells can correlate with reduced endogenous MUC5AC RNA and reduced GFP RNA (from the MUC5AC/GFP constract) to complete validation of hCLCAl.
Example 9: Identification of Potential Target Sites in Human CLCAl RNA
The sequence of human CLCAl is screened for accessible sites using a computer-folding algorithm. Regions of the RNA are identified that do not form secondary folding structures. These regions contain potential ribozyme and/or antisense binding/cleavage sites. The sequences of these binding/cleavage sites are shown in Tables III-IX.
Example 10: Selection of Enzymatic Nucleic Acid Cleavage Sites in Human CLCAl RNA
Ribozyme target sites are chosen by analyzing sequences of Human CLCAl (GenBank accession numbers: NM_001285 and AF039400) and prioritizing the sites on the basis of folding. Ribozymes are designed that could bind each target and are individually analyzed by computer folding (Christoffersen et al, 1994 J. Mol. Struc. Theochem, 311, 273; Jaeger et al, 1989, Proc. Natl. Acad. Sci. USA, 86, 7706) to assess whether the ribozyme sequences fold into the appropriate secondary structure. Those ribozymes with unfavorable intramolecular interactions between the binding arms and the catalytic core are eliminated from consideration. As noted below, varying.binding arm lengths can be chosen to optimize activity. Generally, at least 5 bases on each arm are able to bind to, or otherwise interact with, the target RNA.
Example 11 : Chemical Synthesis and Purification of Ribozymes and Antisense for Efficient Cleavage and/or blocking of CLCAl RNA
Ribozymes and antisense constructs are designed to anneal to various sites in the RNA message. The binding arms of the ribozymes are complementary to the target site sequences described above, while the antisense constructs are fully complimentary to the target site sequences described above. The ribozymes and antisense constructs were chemically synthesized. The method of synthesis used followed the procedure for normal RNA synthesis as described above and in Usman et al, (1987 J. Am. Chem. Soc, 109, 7845), Scaringe et al, (1990 Nucleic Acids Res., 18, 5433) and Wincott et al, supra, and made use of common nucleic acid protecting and coupling groups, such as dimethoxytrityl at the 5'-end, and phosphoramidites at the 3'-end. The average stepwise coupling yields were typically >98%. Ribozymes and antisense constructs are also synthesized from DNA templates using bacteriophage T7 RNA polymerase (Milligan and Uhlenbeck, 1989, Methods Enzymol 180, 51). Ribozymes and antisense constructs are purified by gel electrophoresis using general methods or are purified by high pressure liquid chromatography (HPLC; see Wincott et al, supra; the totality of which is hereby incoφorated herein by reference) and are resuspended in water. The sequences of the chemically synthesized ribozymes and antisense constructs used in this study are shown below in Table III-IX.
Indications
Particular conditions and disease states that can be associated with CLCAl expression modulation include but are not limited to Chronic Obstructive Pulmonary Disease (COPD), chronic bronchitis, asthma, cystic fibrosis, obstructive bowel syndrome, and any other diseases or conditions that are related to or will respond to the levels of CLCAl in a cell or tissue, alone or in combination with other therapies.
The present body of knowledge in CLCAl research indicates the need for methods to assay CLCAl activity and for compounds that can regulate CLCAl expression for research, diagnostic, and therapeutic use.
The nucleic acid molecules of the present invention may also be administered to a patient in combination with other therapeutic compounds to increase the overall therapeutic effect. The use of multiple compounds to treat an indication may increase the beneficial effects while reducing the presence of side effects. Oxygen therapy, bronchodilators, corticosteroids, antibacterials, vaccinations, acetylcysteine, mucokinetic agents, and DNase (Pulmozyme), are non-limiting examples of methods and/or treatments that can be used in combination with nucleic acid molecules of the invention. Those skilled in the art will recognize that other drug compounds and therapies can be similarly and readily combined with the nucleic acid molecules of the instant invention (e.g. ribozymes and antisense molecules) and are, therefore, within the scope of the instant invention.
Cell Culture
The cell culture system described in Example 8 can be used to evaluate nucleic acid molecules of the invention for efficacy in CLCAl and mucin modulation. Animal Models
Numerous reports can be found which describe animal models relevant to disease states such as COPD and cystic fibrosis. These models can be used to determine efficacy of the nucleic acid molecules of the instant invention targeting such disease states or conditions. Animal models for chronic pulmonary disease (COPD) are described by Shapiro, 2000, Am. J. Respir. Cell Mol Biol, 22(1), 4-7; Hogg, 1998, Ika Daigaku Zasshi, 56(3), 429-432; and Garssen et al, 1997, Inhalation Toxicol, 9(6), 581-599. Animal models for cystic fibrosis are described by Kent et al, 1997, J. Clin. Invest, 100(12), 3060-3069; Hill et al, 1991, 62(1), 113-122; Grubb and Gabriel, 1997, Am. J. Physiol, 272, G258-G266; Rozmahel, 1996, From: Diss. Abstr. Int. B 1997, 57(8), 4863; Van Doominck et al, 1995, EMBOJ., 14(18), 4403-11; and Zeiher et al, 1995, J. Clin. Invest, 96(4), 2051-64.
Diagnostic uses
The nucleic acid molecules of this invention (e.g., ribozymes) may be used as diagnostic tools to examine genetic drift and mutations within diseased cells or to detect the presence of CLCAl RNA in a cell. The close relationship between ribozyme activity and the structure of the target RNA allows the detection of mutations in any region of the molecule which alters the base-pairing and three- dimensional stracture of the target RNA. By using multiple ribozymes described in this invention, one may map nucleotide changes which are important to RNA structure and function in vitro, as well as in cells and tissues. Cleavage of target RNAs with ribozymes may be used to inhibit gene expression and define the role (essentially) of specified gene products in the progression of disease. In this manner, other genetic targets may be defined as important mediators of the disease. These experiments will lead to better treatment of the disease progression by affording the possibility of combinational therapies (e.g., multiple ribozymes targeted to different genes, ribozymes coupled with known small molecule inhibitors, or intermittent treatment with combinations of ribozymes and/or other chemical or biological molecules). Other in vitro uses of ribozymes of this invention are well known in the art, and include detection of the presence of mRNAs associated with CLCAl -related condition. Such RNA is detected by determining the presence of a cleavage product after treatment with a ribozyme using standard methodology.
In a specific example, ribozymes which can cleave only wild-type or mutant forms of the target RNA are used for the assay. The first ribozyme is used to identify wild-type RNA present in the sample and the second ribozyme will be used to identify mutant RNA in the sample. As reaction controls, synthetic substrates of both wild-type and mutant RNA will be cleaved by both ribozymes to demonstrate the relative ribozyme efficiencies in the reactions and the absence of cleavage of the "non-targeted" RNA species. The cleavage products from the synthetic substrates will also serve to generate size markers for the analysis of wild-type and mutant RNAs in the sample population. Thus, each analysis can require two ribozymes, two substrates and one unknown sample, which will be combined into six reactions. The presence of cleavage products will be determined using an RNAse protection assay so that full-length and cleavage fragments of each RNA can be analyzed in one lane of a polyacrylamide gel. It is not absolutely required to quantify the results to gain insight into the expression of mutant RNAs and putative risk of the desired phenotypic changes in target cells. The expression of mRNA whose protein product is implicated in the development of the phenotype (i.e., CLCAl) is adequate to establish risk. If probes of comparable specific activity are used for both transcripts, then a qualitative comparison of RNA levels will be adequate and will decrease the cost of the initial diagnosis. Higher mutant form to wild-type ratios will be correlated with higher risk whether RNA levels are compared qualitatively or quantitatively.
Additional Uses
Potential usefulness of sequence-specific enzymatic nucleic acid molecules of the instant invention might have many of the same applications for the study of RNA that DNA restriction endonucleases have for the study of DNA (Nathans et al, 1975 Ann. Rev. Biochem. 44:273). For example, the pattern of restriction fragments could be used to establish sequence relationships between two related RNAs, and large RNAs could be specifically cleaved to fragments of a size more useful for study. The ability to engineer sequence specificity of the enzymatic nucleic acid molecule is ideal for cleavage of RNAs of unknown sequence. Applicant describes the use of nucleic acid molecules to down-regulate gene expression of target genes in bacterial, microbial, fungal, viral, and eukaryotic systems including plant, or mammalian cells.
All patents and publications mentioned in the specification are indicative of the levels of skill of those skilled in the art to which the invention pertains. All references cited in this disclosure are incoφorated by reference to the same extent as if each reference had been incoφorated by reference in its entirety individually.
One skilled in the art would readily appreciate that the present invention is well adapted to carry out the objects and obtain the ends and advantages mentioned, as well as those inherent therein. The methods and compositions described herein as presently representative of preferred embodiments are exemplary and are not intended as limitations on the scope of the invention. Changes therein and other uses will occur to those skilled in the art, which are encompassed within the spirit of the invention, are defined by the scope of the claims.
It will be readily apparent to one skilled in the art that varying substitutions and modifications may be made to the invention disclosed herein without departing from the scope and spirit of the invention. Thus, such additional embodiments are within the scope of the present invention and the following claims.
The invention illustratively described herein suitably may be practiced in the absence of any element or elements, limitation or limitations which is not specifically disclosed herein. Thus, for example, in each instance herein any of the terms "comprising", "consisting essentially of and "consisting of may be replaced with either of the other two terms. The terms and expressions which have been employed are used as terms of description and not of limitation, and there is no intention that in the use of such terms and expressions of excluding any equivalents of the features shown and described or portions thereof, but it is recognized that various modifications are possible within the scope of the invention claimed. Thus, it should be understood that although the present invention has been specifically disclosed by preferred embodiments, optional features, modification and variation of the concepts herein disclosed may be resorted to by those skilled in the art, and that such modifications and variations are considered to be within the scope of this invention as defined by the description and the appended claims.
In addition, where features or aspects of the invention are described in terms of Markush groups or other grouping of alternatives, those skilled in the art will recognize that the invention is also thereby described in terms of any individual member or subgroup of members of the Markush group or other group.
Other embodiments are within the following claims. TABLE I
Characteristics of naturally occurring ribozymes Group I Introns
Size: -150 to >1000 nucleotides.
• Requires a U in the target sequence immediately 5' of the cleavage site.
• Binds 4-6 nucleotides at the 5'-side of the cleavage site.
• Reaction mechanism: attack by the 3' -OH of guanosine to generate cleavage products with 3'-OH and 5'-guanosine.
• Additional protein cofactors required in some cases to help folding and maintainance of the active structure.
• Over 300 known members of this class. Found as an intervening sequence in Tetrahymena thermophila rRNA, fungal mitochondria, chloroplasts, phage T4, blue-green algae, and others.
• Major structural features largely established through phylogenetic comparisons, mutagenesis, and biochemical studies p,u].
• Complete kinetic framework established for one ribozyme ["VW1]*
• Studies of ribozyme folding and substrate docking underway [vii,viii / i ]'
• Chemical modification investigation of important residues well established
[x,xil-
• The small (4-6 nt) binding site may make this ribozyme too non-specific for targeted RNA cleavage, however, the Tetrahymena group I intron has been used to repair a "defective" D -galactosidase message by the ligation of new D- galactosidase sequences onto the defective message [xu].
RNAse P RNA (Ml RNA)
• Size: ~290 to 400 nucleotides.
• RNA portion of a ubiquitous ribonucleoprotein enzyme.
• Cleaves tRNA precursors to form mature tRNA [xiii].
• Reaction mechanism: possible attack by M -OH to generate cleavage products with 3' -OH and 5'-phosphate.
• RNAse P is found throughout the prokaryotes and eukaryotes. The RNA subunit has been sequenced from bacteria, yeast, rodents, and primates.
• Recruitment of endogenous RNAse P for therapeutic applications is possible through hybridization of an External Guide Sequence (EGS) to the target RNA xiv xvl
• Important phosphate and 2' OH contacts recently identified [OTi,xvii] Group II Introns
• Size: >1000 nucleotides.
• Trans cleavage of target RNAs recently demonstrated [xria,xix]. • Sequence requirements not fully determined.
• Reaction mechanism: 2' -OH of an internal adenosine generates cleavage products with 3' -OH and a "lariat" RNA containing a 3'-5' and a 2'-5' branch point.
• Only natural ribozyme with demonstrated participation in DNA cleavage [xx,xxi] in addition to RNA cleavage and ligation.
• Major structural features largely established through phylogenetic comparisons
["xxiil .
• Important 2' OH contacts beginning to be identified [xxu j
• Kinetic framework under development [xxiv]
Neurospora VS RNA
• Size: ~144 nucleotides.
• Trans cleavage of hairpin target RNAs recently demonstrated [xxv].
• Sequence requirements not fully determined.
• Reaction mechanism: attack by 2'-OH 5' to the scissile bond to generate cleavage products with 2',3'-cyclic phosphate and 5'-OH ends.
• Binding sites and structural requirements not fully determined.
• Only 1 known member of this class. Found in Neurospora VS RNA.
Hammerhead Ribozyme
(see text for references)
• Size: ~13 to 40 nucleotides.
• Requires the target sequence UH immediately 5' of the cleavage site.
• Binds a variable number nucleotides on both sides of the cleavage site.
• Reaction mechanism: attack by 2'-OH 5' to the scissile bond to generate cleavage products with 2',3'-cyclic phosphate and 5'-OH ends.
• 14 known members of this class. Found in a number of plant pathogens (virusoids) that use RNA as the infectious agent.
• Essential structural features largely defined, including 2 crystal structures
[xxvi xxviil
• Minimal-l-igat-ion-aetiv-ity demonstrated (for engineering through in vitro selection) [xxviii]
• Complete kinetic framework established for two or more ribozymes [χχiχ].
• Chemical modification investigation of important residues well established
[XXX].
Hairpin Ribozyme
• Size: ~50 nucleotides.
• Requires the target sequence GUC immediately 3' of the cleavage site.
• Binds 4-6 nucleotides at the 5'-side of the cleavage site and a variable number to the 3'-side of the cleavage site.
• Reaction mechanism: attack by 2'-OH 5' to the scissile bond to generate cleavage products with 2',3' -cyclic phosphate and 5' -OH ends. • 3 known members of this class. Found in three plant pathogen (satellite RNAs of the tobacco ringspot virus, arabis mosaic virus and chicory yellow mottle virus) which uses RNA as the infectious agent.
• Essential structural features largely defined [χχχi χχa χ χiii.Modv]
• Ligation activity (in addition to cleavage activity) makes ribozyme amenable to engineering through in vitro selection [x xv]
• Complete kinetic framework established for one ribozyme [xxxvi].
• Chemical modification investigation of important residues begun [χχ ii, χ iu]#
Hepatitis Delta Virus (HDV) Ribozyme
• Size: ~60 nucleotides.
• Trans cleavage of target RNAs demonstrated [xχχix].
• Binding sites and structural requirements not fully determined, although no sequences 5' of cleavage site are required. Folded ribozyme contains a pseudoknot structure [x1].
• Reaction mechanism: attack by 2'-OH 5' to the scissile bond to generate cleavage products with 2',3'-cyclic phosphate and 5' -OH ends.
• Only 2 known members of this class. Found in human HDV.
• Circular form of HDV is active and shows increased nuclease stability [xli]
> . Michel, Francois; Westhof, Eric. Slippery substrates. Nat. Struct. Biol. (1994), 1(1), 5-7. fi . Lisacek, Frederique; Diaz, Yolande; Michel, Francois. Automatic identification of group I intron cores in genomic DNA sequences. J. Mol. Biol. (1994), 235(4), 1206-17.
Hi . Herschlag, Daniel; Cech, Thomas R.. Catalysis of RNA cleavage by the Tetrahymena thermophila ribozyme. 1. Kinetic description of the reaction of an RNA substrate complementary to the active site. Biochemistry (1990), 29(44), 10159-71. iv . Herschlag, Daniel; Cech, Thomas R.. Catalysis of RNA cleavage by the Tetrahymena thermophila ribozyme. 2. Kinetic description of the reaction of an RNA substrate that forms a mismatch at the active site. Biochemistry (1990), 29(44), 10172-80. v . Knitt, Deborah S.; Herschlag, Daniel. pH Dependencies of the Tetrahymena Ribozyme Reveal an Unconventional Origin of an Apparent pKa. Biochemistry (1996), 35(5), 1560-70. vi . Bevilacqua, Philip C; Sugimoto, Naoki; Turner, Douglas H.. A mechanistic framework for the second step of splicing catalyzed by the Tetrahymena ribozyme. Biochemistry (1996), 35(2), 648-58. vii . Li, Yi; Bevilacqua, Philip C; Ma thews, David; Turner, Douglas H.. Thermodynamic and activation parameters for binding of a pyrene-labeled substrate by the Tetrahymena ribozyme: docking is not diffusion-controlled and is driven by a favorable entropy change. Biochemistry (1995),
34(44), 14394-9. v . Banerjee, Aloke Raj; Turner, Douglas H„ The time dependence of chemical modification reveals slow steps in the folding of a group I ribozyme. Biochemistry (1995), 34(19), 6504-12.
■" . Zarrinkar, Patrick P.; Williamson, James R.. The P9.1-P9.2 peripheral extension helps guide folding of the Tetrahymena ribozyme. Nucleic Acids Res. (1996), 24(5), 854-8. x . Strobel, Scott A.; Cech, Thomas R.. Minor groove recognition of the conserved G.cntdot.U pair at the Tetrahymena ribozyme reaction site. Science (Washington, D. C.) (1995), 267(5198), 675-9. n . Strobel, Scott A.; Cech, Thomas R.. Exocyclic Amine of the Conserved G.cntdotU Pair at the
Cleavage Site of the Tetrahymena Ribozyme Contributes to 5'-Splice Site Selection and Transition
State Stabilization. Biochemistry (1996), 35(4), 1201-11.
*&. Sullenger, Bruce A.; Cech, Thomas R.. Ribozyme-mediated repair of defective mRNA by targeted trans-splicing. Nature (London) (1994), 371(6498), 619-22. . Robertson, H.D.; Altman, S.; Smith, J.D. J. Biol. Chem., 247, 5243-5251 (1972). xiv. Forster, Anthony C; Altman, Sidney. External guide sequences for an RNA enzyme. Science (Washington, D. C, 1883-) (1990), 249(4970), 783-6. xv. Yuan, Y.; Hwang, E. S.; Altman, S. Targeted cleavage of mRNA by human RNase P. Proc.
Natl. Acad. Sci. USA (1992) 89, 8006-10. xvi . Harris, Michael E.; Pace, Norman R.. Identification of phosphates involved in catalysis by the ribozyme RNase P RNA. RNA (1995), 1(2), 210-18. x™ . Pan, Tao; Loria, Andrew; Zhong, Kun. Probing of tertiary interactions in RNA: 2'-hydroxyl- base contacts between the RNase P RNA and pre-tRNA. Proc. Natl. Acad. Sci. U. S. A. (1995), 92(26),
12510-14. xvifi Pyle, Anna Marie; Green, Justin B.. Building a Kinetic Framework for Group II Intron
Ribozyme Activity: Quantitation of Interdomain Binding and Reaction Rate. Biochemistry (1994),
33(9), 2716-25. ix . Michels, William J. Jr.; Pyle, Anna Marie. Conversion of a Group II Intron into a New
Multiple-Turnover Ribozyme that Selectively Cleaves Oligonucleotides: Elucidation of Reaction
Mechanism and Structure/ Function Relationships. Biochemistry (1995), 34(9), 2965-77. x . Zimmerly, Steven; Guo, Huatao; Eskes, Robert; Yang, Jian; Perlman, Philip S.; Lambowitz,
Alan M.. A group II intron RNA is a catalytic component of a DNA endonuclease involved in intron mobility. Cell (Cambridge, Mass.) (1995), 83(4), 529-38. xxi . Griffin, Edmund A., Jr.; Qin, Zhifeng; Michels, Williams J., Jr.; Pyle, Anna Marie. Group II intron ribozymes that cleave DNA and RNA linkages with similar efficiency, and lack contacts with substrate 2'-hydroxyl groups. Chem. Biol. (1995), 2(11), 761-70. x ϋ . Michel, Francois; Ferat, Jean Luc. Structure and activities of group II introns. Annu. Rev.
Biochem. (1995), 64, 435-61. xxϋi . Abramovitz, Dana L.; Friedman, Richard A.; Pyle, Anna Marie. Catalytic role of 2'-hydroxyl groups within a group II intron active site. Science (Washington, D. C.) (1996), 271(5254), 1410-13. xxiv . Daniels, Danette L.; Michels, William J., Jr.; Pyle, Anna Marie. Two competing pathways for self-splicing by group II introns: a quantitative analysis of in vitro reaction rates and products. J. Mol.
Biol. (1996), 256(1), 31-49. xxv . Guo, Hans C. T.; Collins, Richard A.. Efficient trans-cleavage of a stem-loop RNA substrate by a ribozyme derived from Neurospora VS RNA. EMBO J. (1995), 14(2), 368-76. xxvi . Scott, W.G., Finch, J.T., Aaron,K. The crystal structure of an all RNA hammerhead ribozyme:Aproposed mechanism for RNA catalytic cleavage. Cell, (1995), 81, 991-1002. xx ii McKay, Structure and function of the hammerhead ribozyme: an unfinished story. RNA,
(1996), 2, 395-403. xxviii . Long, D., Uhlenbeck, O., Hertel, K. Ligation with hammerhead ribozymes. US Patent No.
5,633,133. xix . Hertel, K.J., Herschlag, D., Uhlenbeck, O. A kinetic and thermodynamic framework for the hammerhead ribozyme reaction. Biochemistry, (1994) 33, 3374-3385. Beigelman, L., et al., Chemical modifications of hammerhead ribozymes. J. Biol. Chem., (1995) 270, 25702-25708. xx . Beigelman, L., et al, Chemical modifications of hammerhead ribozymes. J. Biol. Chem., (1995)
270, 25702-25708. xxxi . Hampel, Arnold; Tritz, Richard; Hicks, Margaret; Cruz, Phillip. 'Hairpin' catalytic RNA model: evidence for helixes and sequence requirement for substrate RNA. Nucleic Acids Res. (1990),
18(2), 299-304. xxxii . Chowrira, Bharat M.; Berzal-Herra z, Alfredo; Burke, John M.. Novel guanosine requirement for catalysis by the hairpin ribozyme. Nature (London) (1991), 354(6351), 320-2. x xui Berzal-Herranz, Alfredo; Joseph, Simpson; Chowrira, Bharat M.; Butcher, Samuel E.; Burke,
John M.. Essential nucleotide sequences and secondary structure elements of the hairpin ribozyme.
EMBO J. (1993), 12(6), 2567-73. xxxiv Joseph, Simpson; Berzal-Herranz, Alfredo; Chowrira, Bharat M.; Butcher, Samuel E..
Substrate selection rules for the hairpin ribozyme determined by in vitro selection, mutation, and analysis of mismatched substrates. Genes Dev. (1993), 7(1), 130-8. xxxv Berzal-Herranz, Alfredo; Joseph, Simpson; Burke, John M.. In vitro selection of active hairpin ribozymes by sequential RNA-catalyzed cleavage and ligation reactions. Genes Dev. (1992), 6(1), 129-
34. x xvi Hegg, Lisa A.; Fedor, Martha J.. Kinetics and Thermodynamics of Intermolecular Catalysis by Hairpin Ribozymes. Biochemistry (1995), 34(48), 15813-28. xx vii Grasby, Jane A.; Mersmann, Karin; Singh, Mohinder; Gait, Michael J.. Purine Functional Groups in Essential Residues of the Hairpin Ribozyme Required for Catalytic Cleavage of RNA. Biochemistry (1995), 34(12), 4068-76. xxxviu Schmidt, Sabine; Beigelman, Leonid; Karpeisky, Alexander; Usman, Nassim; Sorensen, Ulrik S.; Gait, Michael J.. Base and sugar requirements for RNA cleavage of essential nucleoside residues in internal loop B of the hairpin ribozyme: implications for secondary structure. Nucleic Acids Res. (1996), 24(4), 573-81. xxxix Perrotta, Anne T.; Been, Michael D.. Cleavage of oligoribonucleotides by a ribozyme derived from the hepatitis .delta, virus RNA sequence. Biochemistry (1992), 31(1), 16-21. l . Perrotta, Anne T.; Been, Michael D.. A pseudoknot-like structure required for efficient self- cleavage of hepatitis delta virus RNA. Nature (London) (1991), 350(6317), 434-6. *& . Puttaraju, M.; Perrotta, Anne T.; Been, Michael D.. A circular trans-acting hepatitis delta virus ribozyme. Nucleic Acids Res. (1993), 21(18), 4253-8.
Table II:
A. 2.5 μmol Synthesis Cycle ABI 394 instrument
Figure imgf000053_0001
Figure imgf000054_0001
C. 0.2 μmol Synthesis Cycle 96 well Instrument
Figure imgf000054_0002
Wait time does not include contact time during delivery.
Table III: Human CLCAl Hammerhead Ribozyme and Target Sequence 249.021
Pos Substrate Seq ID Ribozyme Rz
No. Seq ID
No.
11 CUAAUGCU U UUGGUACA 1 UGUACCAA CUGAUGAG GCCGUUAGGC CGAA AGCAUUAG 2190
12 UAAUGCUU u UGGUΆCΆA 2 UUGUACCA CUGAUGAG GCCGUUAGGC CGAA AAGCAUUA 2191
13 AAUGCUUU U GGUACAAA 3 UUUGUACC CUGAUGAG GCCGUUAGGC CGAA. AAAGCAUU 2192
17 CUUUUGGU A CAAAUGGA 4 UCCAUUUG CUGAUGAG GCCGUUAGGC CGAA ACCAAAAG 2193
34 UGUGGAAU A UAAUUGAA 5 UUCAAUUA CUGAUGAG GCCGUUAGGC CGAA AUUCCACA 2194
36 UGGAAUAU A AUUGAAUA 6 UAUUCAAU CUGAUGAG GCCGUUAGGC CGAA AUAUUCCA 2195
39 AAUAUAAU U GAAUAUUU 7 AAAUAUUC CUGAUGAG GCCGUUAGGC CGAA AUUAUAUU 2196
44 AAUUGAAU A UUUUCUUG 8 CAAGAAAA CUGAUGAG GCCGUUAGGC CGAA AUUCAAUU 2197
46 UUGAAUAU U UUCUUGUU 9 AACAAGAA CUGAUGAG GCCGUUAGGC CGAA AUAUUCAA 2198
47 UGAAUAUU U UCUUGUUU 10 AAACAAGA CUGAUGAG GCCGUUAGGC CGAA AAUAUUCA 2199
48 GAAUAUUU U CUUGUUUA 11 UAAACAAG CUGAUGAG GCCGUUAGGC CGAA AAAUAUUC 2200
49 AAUAUUUU C UUGUUUAA 12 UUAAACAA CUGAUGAG GCCGUUAGGC CGAA AAAAUAUU 2201
51 UAUUUUCU U GUUUAAGG 13 CCUUAAAC CUGAUGAG GCCGUUAGGC CGAA AGAAAAUA 2202
54 UUUCUUGU U UAAGGGGA 14 UCCCCUUA CUGAUGAG GCCGUUAGGC CGAA ACAAGAAA 2203
55 UUCUUGUU U AAGGGGAG 15 CUCCCCUU CUGAUGAG GCCGUUAGGC CGAA AACAAGAA 2204
56 UCUUGUUU A AGGGGAGC 16 GCUCCCCU CUGAUGAG GCCGUUAGGC CGAA AAACAAGA 2205
77 AGAGGUGU U GAGGUUAU 17 AUAACCUC CUGAUGAG GCCGUUAGGC CGAA ACACCUCU 2206
83 GUUGAGGU U AUGUCAAG 18 CUUGACAU CUGAUGAG GCCGUUAGGC CGAA ACCUCAAC 2207
84 UUGAGGUU A UGUCAAGC 19 GCUUGACA CUGAUGAG GCCGUUAGGC CGAA AACCUCAA 2208
88 GGUUAUGU C AAGCAUCU 20 AGAUGCUU CUGAUGAG GCCGUUAGGC CGAA ACAUAACC 2209
95 UCAAGCAU C UGGCACAG 21 CUGUGCCA CUGAUGAG GCCGUUAGGC CGAA AUGCUUGA 2210
122 AUGGAAAU A UUUACAAG 22 CUUGUAAA CUGAUGAG GCCGUUAGGC CGAA AUUUCCAU 2211
124 GGAAAUAU U UACAAGUA 23 UACUUGUA CUGAUGAG GCCGUUAGGC CGAA AUAUUUCC 2212
125 GAAAUAUU U ACAAGUAC 24 GUACUUGU CUGAUGAG GCCGUUAGGC CGAA AAUAUUUC 2213
126 AAAUAUUU A CAAGUACG 25 CGUACUUG CUGAUGAG GCCGUUAGGC CGAA AAAUAUUU 2214
132 UUACAAGU A CGCAAUUU 26 AAAUUGCG CUGAUGAG GCCGUUAGGC CGAA ACUUGUAA 2215
139 UACGCAAU U UGAGACUA 27 UAGUCUCA CUGAUGAG GCCGUUAGGC CGAA AUUGCGUA 2216
140 ACGCAAUU U GAGACUAA 28 UUAGUCUC CUGAUGAG GCCGUUAGGC CGAA AAUUGCGU 2217
147 UUGAGACU A AGAUAUUG 29 CAAUAUCU CUGAUGAG GCCGUUAGGC CGAA AGUCUCAA 2218
152 ACUAAGAU A UUGUUAUC 30 GAUAACAA CUGAUGAG GCCGUUAGGC CGAA AUCUUAGU 2219
154 UAAGAUAU U GUUAUCAU 31 ' AUGAUAAC CUGAUGAG GCCGUUAGGC CGAA AUAUCUUA 2220
157 GAUAUUGU U AUCAUUCU 32 AGAAUGAU CUGAUGAG GCCGUUAGGC CGAA ACAAUAUC 2221
158 AUAUUGUU A UCAUUCUC 33 GAGAAUGA CUGAUGAG GCCGUUAGGC CGAA AACAAUAU 2222
160 AUUGUUAU C AUUCUCCU 34 AGGAGAAU CUGAUGAG GCCGUUAGGC CGAA AUAACAAU 2223
163 GUUAUCAU U CUCCUAUU 35 AAUAGGAG CUGAUGAG GCCGUUAGGC CGAA AUGAUAAC 2224
164 UUAUCAUU C UCCUAUUG 36 CAAUAGGA CUGAUGAG GCCGUUAGGC CGAA AAUGAUAA 2225
166 AUCAUUCU C CUAUUGAA 37 UUCAAUAG CUGAUGAG GCCGUUAGGC CGAA AGAAUGAU 2226
169 AUUCUCCU A UUGAAGAC 38 GUCUUCAA CUGAUGAG GCCGUUAGGC CGAA AGGAGAAU 2227
171 UCUCCUAU U GAAGACAA 39 UUGUCUUC CUGAUGAG GCCGUUAGGC CGAA AUAGGAGA 2228
187 AGAGCAAU A GUAAAACA 40 UGUUUUAC CUGAUGAG GCCGUUAGGC CGAA AUUGCUCU 2229
190 GCAAUAGU A AAACACAU 41 AUGUGUUU CUGAUGAG GCCGUUAGGC CGAA ACUAUUGC 2230
199 AAACACAU C AGGUCAGG 42 CCUGACCU CUGAUGAG GCCGUUAGGC CGAA AUGUGUUU 2231
204 CAUCAGGU C AGGGGGUU 43 AACCCCCU CUGAUGAG GCCGUUAGGC CGAA ACCUGAUG 2232
212 CAGGGGGU U AAAGACCU 44 AGGUCUUU CUGAUGAG GCCGUUAGGC CGAA ACCCCCUG 2233
213 AGGGGGUU A AAGACCUG 45 CAGGUCUU CUGAUGAG GCCGUUAGGC CGAA AACCCCCU 2234
226 CCUGUGAU A AACCACUU 46 AAGUGGUU CUGAUGAG GCCGUUAGGC CGAA AUCACAGG 2235
234 AAACCACU U CCGAUAAG 47 CUUAUCGG CUGAUGAG GCCGUUAGGC CGAA AGUGGUTJU 2236
235 AACCACUU C CGAUAAGU 48 ACUUAUCG CUGAUGAG GCCGUUAGGC CGAA AAGUGGUU 2237
240 CUUCCGAU A AGUUGGAA 49 UUCCAACU CUGAUGAG GCCGUUAGGC CGAA AUCGGAAG 2238
244 CGAUAAGU U GGAAACGU 50 ACGUUUCC CUGAUGAG GCCGUUAGGC CGAA ACUUAUCG 2239
257 ACGUGUGU C UAUAUUUU 51 AAAAUAUA CUGAUGAG GCCGUUAGGC CGAA ACACACGU 2240
259 GUGUGUCU A UAUUUUCA 52 UGAAAAUA CUGAUGAG GCCGUUAGGC CGAA AGACACAC 2241 261 GUGUCUAU A UUUUCAUA 53 UAUGAAAA CUGAUGAG GCCGUUAGGC CGAA AUAGACAC 2242
263 GUCUAUAU U UUCAUAUC 54 GAUAUGAA CUGAUGAG GCCGUUAGGC CGAA AUAUAGAC 2243
264 UCUAUAUϋ U UCAUAUCU 55 AGAUAUGA CUGAUGAG GCCGUUAGGC CGAA AAUAUAGA 2244
265 CUAUAUUU U CAUAUCUG 56 CAGAUAUG CUGAUGAG GCCGUUAGGC CGAA AAAUAUAG 2245
266 UAUAUUUU C AUAUCUGU 57 ACAGAUAU CUGAUGAG GCCGUUAGGC CGAA AAAAUAUA 2246
269 AUUUUCAU A UCUGUAUA 58 UAUACAGA CUGAUGAG GCCGUUAGGC CGAA AUGAAAAU 2247
271 UUUCAUAU C UGUAUAUA 59 UAUAUACA CUGAUGAG GCCGUUAGGC CGAA AUAUGAAA 2248
275 AUAUCUGU A UAUAUAUA 60 UAUAUAUA CUGAUGAG GCCGUUAGGC CGAA ACAGAUAU 2249
277 AUCUGUAU A UAUAUAAU 61 AUUAUAUA CUGAUGAG GCCGUUAGGC CGAA AUACAGAU 2250
279 CUGUAUAU A UAUAAUGG 62 CCAUUAUA CUGAUGAG GCCGUUAGGC CGAA AUAUACAG 2251
281 GUAUAUAU A UAAUGGUA 63 UACCAUUA CUGAUGAG GCCGUUAGGC CGAA AUAUAUAC 2252
283 AUAUAUAU A AUGGUAAA 64 UUUACCAU CUGAUGAG GCCGUUAGGC CGAA AUAUAUAU 2253
289 AUAAUGGU A AAGAAAGA 65 UCUUUCUU CUGAUGAG GCCGUUAGGC CGAA ACCAUUAU 2254
303 AGACACCU U CGUAACCC 66 GGGUUACG CUGAUGAG GCCGUUAGGC CGAA AGGUGUCU 2255
304 GACACCUU C GUAACCCG 67 CGGGUUAC CUGAUGAG GCCGUUAGGC CGAA AAGGUGUC 2256
307 ACCUUCGU A ACCCGCAU 68 AUGCGGGU CUGAUGAG GCCGUUAGGC CGAA ACGAAGGU 2257
316 ACCCGCAU U UUCCAAAG 69 CUUUGGAA CUGAUGAG GCCGUUAGGC CGAA AUGCGGGU 2258
317 CCCGCAUU U UCCAAAGA 70 UCUUUGGA CUGAUGAG GCCGUUAGGC CGAA AAUGCGGG 2259
318 CCGCAUUU U CCAAAGAG 71 CUCUUUGG CUGAUGAG GCCGUUAGGC CGAA AAAUGCGG 2260
319 CGCAUUUU C CAAAGAGA 72 UCUCUUUG CUGAUGAG GCCGUUAGGC CGAA AAAAUGCG 2261
333 AGAGGAAU C ACAGGGAG 73 CUCCCUGU CUGAUGAG GCCGUUAGGC CGAA AUUCCUCU 2262
346 GGAGAUGU A CAGCAAUG 74 CAUUGCUG CUGAUGAG GCCGUUAGGC CGAA ACAUCUCC 2263
362 GGGGCCAU U UAAGAGUU 75 AACUCUUA CUGAUGAG GCCGUUAGGC CGAA AUGGCCCC 2264
363 GGGCCAUU U AAGAGUUC 76 GAACUCUU CUGAUGAG GCCGUUAGGC CGAA AAUGGCCC 2265
364 GGCCAUUU A AGAGUUCU 77 AGAACUCU CUGAUGAG GCCGUUAGGC CGAA AAAUGGCC 2266
370 UUAAGAGU U CUGUGUUC 78 GAACACAG CUGAUGAG GCCGUUAGGC CGAA ACUCUUAA 2267
371 UAAGAGUU C UGUGUUCA 79 UGAACACA CUGAUGAG GCCGUUAGGC CGAA AACUCUUA 2268
377 UUCUGUGU U CAUCUUGA 80 UCAAGAUG CUGAUGAG GCCGUUAGGC CGAA ACACAGAA 2269
378 UCUGUGUU C AUCUUGAU 81 AUCAAGAU CUGAUGAG GCCGUUAGGC CGAA AACACAGA 2270
381 GUGUUCAU C UUGAUUCU 82 AGAAUCAA CUGAUGAG GCCGUUAGGC CGAA AUGAACAC 2271
383 GUUCAUCU u GAUUCUUC 83 GAAGAAUC CUGAUGAG GCCGUUAGGC CGAA AGAUGAAC 2272
387 AUCUUGAU u CUUCACCU 84 AGGUGAAG CUGAUGAG GCCGUUAGGC CGAA AUCAAGAU 2273
388 UCUUGAUU C UUCACCUU 85 AAGGUGAA CUGAUGAG GCCGUUAGGC CGAA AAUCAAGA 2274
390 UUGAUUCU u CACCUUCU 86 AGAAGGUG CUGAUGAG GCCGUUAGGC CGAA AGAAUCAA 2275
391 UGAUUCUU C ACCUUCUA 87 UAGAAGGU CUGAUGAG GCCGUUAGGC CGAA AAGAAUCA 2276
396 CUUCACCU u CUAGAAGG 88 CCUUCUAG CUGAUGAG GCCGUUAGGC CGAA AGGUGAAG 2277
397 UUCACCUU C UAGAAGGG 89 CCCUUCUA CUGAUGAG GCCGUUAGGC CGAA AAGGUGAA 2278
399 CACCUUCU A GAAGGGGC 90 GCCCCUUC CUGAUGAG GCCGUUAGGC CGAA AGAAGGUG 2279
415 CCCUGAGU A AUUCACUC 91 GAGUGAAU CUGAUGAG GCCGUUAGGC CGAA ACUCAGGG 2280
418 UGAGUAAU U CACUCAUU 92 AAUGAGUG CUGAUGAG GCCGUUAGGC CGAA AUUACUCA 2281
419 GAGUAAUU C ACUCAUUC 93 GAAUGAGU CUGAUGAG GCCGUUAGGC CGAA AAUUACUC 2282
423 AAUUCACU C AUUCAGCU 94 AGCUGAAU CUGAUGAG GCCGUUAGGC CGAA AGUGAAUU 2283
426 UCACUCAU U CAGCUGAA 95 UUCAGCUG CUGAUGAG GCCGUUAGGC CGAA AUGAGUGA 2284
427 CACUCAUU C AGCUGAAC 96 GUUCAGCU CUGAUGAG GCCGUUAGGC CGAA AAUGAGUG 2285
446 CAAUGGCU A UGAAGGCA 97 UGCCUUCA CUGAUGAG GCCGUUAGGC CGAA AGCCAUUG 2286
456 GAAGGCAU U GUCGUUGC 98 GCAACGAC CUGAUGAG GCCGUUAGGC CGAA AUGCCUUC 2287
459 GGCAUUGU C GUUGCAAU 99 AUUGCAAC CUGAUGAG GCCGUUAGGC CGAA ACAAUGCC 2288
462 AUUGUCGU U GCAAUCGA 100 UCGAUUGC CUGAUGAG GCCGUUAGGC CGAA ACGACAAU 2289
468 GUUGCAAU C GACCCCAA 101 UUGGGGUC CUGAUGAG GCCGUUAGGC CGAA AUUGCAAC 2290
498 GAAACACU C AUUCAACA 102 UGUUGAAU CUGAUGAG GCCGUUAGGC CGAA AGUGUUUC 2291
501 ACACUCAU u CAACAAAU 103 AUUUGUUG CUGAUGAG GCCGUUAGGC CGAA AUGAGUGU 2292
502 CACUCAUU c AACAAAUA 104 UAUUUGUU CUGAUGAG GCCGUUAGGC CGAA AAUGAGUG 2293
510 CAACAAAU A AAGGACAU 105 AUGUCCUU CUGAUGAG GCCGUUAGGC CGAA AUUUGUUG 2294
533 CCAGGCAU c UCUGUAUC 106 GAUACAGA CUGAUGAG GCCGUUAGGC CGAA AUGCCUGG 2295
535 AGGCAUCU c UGUAUCUG 107 CAGAUACA CUGAUGAG GCCGUUAGGC CGAA AGAUGCCU 2296
539 AUCUCUGU A UCUGUUUG 108 CAAACAGA CUGAUGAG GCCGUUAGGC CGAA ACAGAGAU 2297 541 CUCUGUAU C UGUUUGAA 109 UUCAAACA CUGAUGAG GCCGUUAGGC CGAA AUACAGAG 2298
545 GUAUCUGU U UGAAGCUA 110 UAGCUUCA CUGAUGAG GCCGUUAGGC CGAA ACAGAUAC 2299
546 UAUCUGUU U GAAGCUAC 111 GUAGCUUC CUGAUGAG GCCGUUAGGC CGAA AACAGAUA 2300
553 UUGAAGCU A CAGGAAAG 112 CUUUCCUG CUGAUGAG GCCGUUAGGC CGAA AGCUUCAA 2301
566 AAAGCGAU U UUAUUUCA 113 UGAAAUAA CUGAUGAG GCCGUUAGGC CGAA AUCGCUUU 2302
567 AAGCGAUU U UAUUUCAA 114 UUGAAAUA CUGAUGAG GCCGUUAGGC CGAA AAUCGCUU 2303
568 AGCGAUUU U AUUUCAAA 115 UUUGAAAU CUGAUGAG GCCGUUAGGC CGAA AAAUCGCU 2304
569 GCGAUUUU A UUUCAAAA 116 UUUUGAAA CUGAUGAG GCCGUUAGGC CGAA AAAAUCGC 2305
571 GAUUUUAU U UCAAAAAU 117 AUUUUUGA CUGAUGAG GCCGUUAGGC CGAA AUAAAAUC 2306
572 AUUUUAUU U CAAAAAUG 118 CAUUUUUG CUGAUGAG GCCGUUAGGC CGAA AAUAAAAU 2307
573 UUUUAUUU C AAAAAUGU 119 ACAUUUUU CUGAUGAG GCCGUUAGGC CGAA AAAUAAAA 2308
582 AAAAAUGU U GCCAUUUU 120 AAAAUGGC CUGAUGAG GCCGUUAGGC CGAA ACAUUUUU 2309
588 GUUGCCAU U UUGAUUCC 121 GGAAUCAA CUGAUGAG GCCGUUAGGC CGAA AUGGCAAC 2310
589 UUGCCAUU U UGAUUCCU 122 AGGAAUCA CUGAUGAG GCCGUUAGGC CGAA AAUGGCAA 2311
590 UGCCAUUϋ U GAUUCCUG 123 CAGGAAUC CUGAUGAG GCCGUUAGGC CGAA AAAUGGCA 2312
594 AUUUUGAU U CCUGAAAC 124 GUUUCAGG CUGAUGAG GCCGUUAGGC CGAA AUCAAAAU 2313
595 UUUUGAUU C CUGAAACA 125 UGUUUCAG CUGAUGAG GCCGUUAGGC CGAA AAUCAAAA 2314
623 GGCUGACU A UGUGAGAC 126 GUCUCACA CUGAUGAG GCCGUUAGGC CGAA AGUCAGCC 2315
639 CCAAAACU U GAGACCUA 127 UAGGUCUC CUGAUGAG GCCGUUAGGC CGAA AGUUUUGG 2316
647 UGAGACCU A CAAAAAUG 128 CAUUUUUG CUGAUGAG GCCGUUAGGC CGAA AGGUCUCA 2317
663 GCUGAUGU U CUGGUUGC 129 GCAACCAG CUGAUGAG GCCGUUAGGC CGAA ACAUCAGC 2318
664 CUGAUGUU C UGGUUGCU 130 AGCAACCA CUGAUGAG GCCGUUAGGC CGAA AACAUCAG 2319
669 GUUCUGGU U GCUGAGUC 131 GACUCAGC CUGAUGAG GCCGUUAGGC CGAA ACCAGAAC 2320
677 UGCUGAGU c UACUCCUC 132 GAGGAGUA CUGAUGAG GCCGUUAGGC CGAA ACUCAGCA 2321
679 CUGAGUCU A CUCCUCCA 133 UGGAGGAG CUGAUGAG GCCGUUAGGC CGAA AGACUCAG 2322
682 AGUCUACU C CUCCAGGU 134 ACCUGGAG CUGAUGAG GCCGUUAGGC CGAA AGUAGACU 2323
685 CUACUCCU c CAGGUAAU 135 AUUACCUG CUGAUGAG GCCGUUAGGC CGAA AGGAGUAG 2324
691 CUCCAGGU A AUGAUGAA 136 UUCAUCAU CUGAUGAG GCCGUUAGGC CGAA ACCUGGAG 2325
704 UGAACCCU A CACUGAGC 137 GCUCAGUG CUGAUGAG GCCGUUAGGC CGAA AGGGUUCA 2326
747 GAAAGGAU C CACCUCAC 138 GUGAGGUG CUGAUGAG GCCGUUAGGC CGAA AUCCUUUC 2327
753 AUCCACCU C ACUCCUGA 139 UCAGGAGU CUGAUGAG GCCGUUAGGC CGAA AGGUGGAU 2328
757 ACCUCACU C CUGAUUUC 140 GAAAUCAG CUGAUGAG GCCGUUAGGC CGAA AGUGAGGU 2329
763 CUCCUGAU U UCAUUGCA 141 UGCAAUGA CUGAUGAG GCCGUUAGGC CGAA AUCAGGAG 2330
764 UCCUGAUU U CAUUGCAG 142 CUGCAAUG CUGAUGAG GCCGUUAGGC CGAA AAUCAGGA 2331
765 CCUGAUUU C AUUGCAGG 143 CCUGCAAU CUGAUGAG GCCGUUAGGC CGAA AAAUCAGG 2332
768 GAUUUCAU u GCAGGAAA 144 UUUCCUGC CUGAUGAG GCCGUUAGGC CGAA AUGAAAUC 2333
782 AAAAAAGU u AGCUGAAU 145 AUUCAGCU CUGAUGAG GCCGUUAGGC CGAA ACUUUUUU 2334
783 AAAAAGUU A GCUGAAUA 146 UAUUCAGC CUGAUGAG GCCGUUAGGC CGAA AACUUUUU 2335
791 AGCUGAAU A UGGACCAC 147 GUGGUCCA CUGAUGAG GCCGUUAGGC CGAA AUUCAGCU 2336
805 CACAAGGU A AGGCAUUU 148 AAAUGCCU CUGAUGAG GCCGUUAGGC CGAA ACCUUGUG 2337
812 UAAGGCAU U UGUCCAUG 149 CAUGGACA CUGAUGAG GCCGUUAGGC CGAA AUGCCUUA 2338
813 AAGGCAUU U GUCCAUGA 150 UCAUGGAC CUGAUGAG GCCGUUAGGC CGAA AAUGCCUU 2339
816 GCAUUUGU C CAUGAGUG 151 CACUCAUG CUGAUGAG GCCGUUAGGC CGAA ACAAAUGC 2340
829 AGUGGGCU C AUCUACGA 152 UCGUAGAU CUGAUGAG GCCGUUAGGC CGAA AGCCCACU 2341
832 GGGCUCAU C UACGAUGG 153 CCAUCGUA CUGAUGAG GCCGUUAGGC CGAA AUGAGCCC 2342
834 GCUCAUCU A CGAUGGGG 154 CCCCAUCG CUGAUGAG GCCGUUAGGC CGAA AGAUGAGC 2343
846 UGGGGAGU A UUUGACGA 155 UCGUCAAA CUGAUGAG GCCGUUAGGC CGAA ACUCCCCA 2344
848 GGGAGUAU U UGACGAGU 156 ACUCGUCA CUGAUGAG GCCGUUAGGC CGAA AUACUCCC 2345
849 GGAGUAUU U GACGAGUA 157 UACUCGUC CUGAUGAG GCCGUUAGGC CGAA AAUACUCC 2346
857 UGACGAGU A CAAUAAUG 158 CAUUAUUG CUGAUGAG GCCGUUAGGC CGAA ACUCGUCA 2347
862 AGUACAAU A AUGAUGAG 159 CUCAUCAU CUGAUGAG GCCGUUAGGC CGAA AUUGUACU 2348
875 UGAGAAAU U CUACUUAU 160 AUAAGUAG CUGAUGAG GCCGUUAGGC CGAA AUUUCUCA 2349
876 GAGAAAUU C UACUUAUC 161 GAUAAGUA CUGAUGAG GCCGUUAGGC CGAA AAUUUCUC 2350
878 GAAAUUCU A CUUAUCCA 162 UGGAUAAG CUGAUGAG GCCGUUAGGC CGAA AGAAUUUC 2351
881 AUUCUACU U AUCCAAUG 163 CAUUGGAU CUGAUGAG GCCGUUAGGC CGAA AGUAGAAU 2352
882 UUCUACUU A UCCAAUGG 164 CCAUUGGA CUGAUGAG GCCGUUAGGC CGAA AAGUAGAA 2353 884 CUACUUAU C CAAUGGAA 165 UUCCAUUG CUGAUGAG GCCGUUAGGC CGAA AUAAGUAG 2354
897 GGAAGAAU A CAAGCAGU 166 ACUGCUUG CUGAUGAG GCCGUUAGGC CGAA AUUCUUCC 2355
906 CAAGCAGU A AGAUGUUC 167 GAACAUCU CUGAUGAG GCCGUUAGGC CGAA ACUGCUUG 2356
913 UAAGAUGU U CAGCAGGU 168 ACCϋGCUG CUGAUGAG GCCGUUAGGC CGAA ACAUCUUA 2357
914 AAGAUGUU C AGCAGGUA 169 UACCUGCU CUGAUGAG GCCGUUAGGC CGAA AACAUCUU 2358
922 CAGCAGGU A UUACUGGU 170 ACCAGUAA CUGAUGAG GCCGUUAGGC CGAA ACCUGCUG 2359
924 GCAGGUAU U ACUGGUAC 171 GUACCAGU CUGAUGAG GCCGUUAGGC CGAA AUACCUGC 2360
925 CAGGUAUU A CUGGUACA 172 UGUACCAG CUGAUGAG GCCGUUAGGC CGAA AAUACCUG 2361
931 UUACUGGU A CAAAUGUA 173 UACAUUUG CUGAUGAG GCCGUUAGGC CGAA ACCAGUAA 2362
939 ACAAAUGU A GUAAAGAA 174 UUCUUUAC CUGAUGAG GCCGUUAGGC CGAA ACAUUUGU 2363
942 AAUGUAGU A AAGAAGUG 175 CACUUCUU CUGAUGAG GCCGUUAGGC CGAA ACUACAUU 2364
952 AGAAGUGU C AGGGAGGC 176 GCCUCCCU CUGAUGAG GCCGUUAGGC CGAA ACACUUCU 2365
967 GCAGCUGU U ACACCAAA 177 UUUGGUGU CUGAUGAG GCCGUUAGGC CGAA ACAGCUGC 2366
968 CAGCUGUU A CACCAAAA 178 UUUUGGUG CUGAUGAG GCCGUUAGGC CGAA AACAGCUG 2367
986 AUGCACAU U CAAUAAAG 179 CUUUAUUG CUGAUGAG GCCGUUAGGC CGAA AUGUGCAU 2368
987 UGCACAUU C AAUAAAGU 180 ACUUUAUU CUGAUGAG GCCGUUAGGC CGAA AAUGUGCA 2369
991 CAUUCAAU A AAGUUACA 181 UGUAACUU CUGAUGAG GCCGUUAGGC CGAA AUUGAAUG 2370
996 AAUAAAGU U ACAGGACU 182 AGUCCUGU CUGAUGAG GCCGUUAGGC CGAA ACUUUAUU 2371
997 AUAAAGUU A CAGGACUC 183 GAGUCCUG CUGAUGAG GCCGUUAGGC CGAA AACUUUAU 2372
1005 ACAGGACU C UAUGAAAA 184 UUUUCAUA CUGAUGAG GCCGUUAGGC CGAA AGUCCUGU 2373
1007 AGGACUCU A UGAAAAAG 185 CUUUUUCA CUGAUGAG GCCGUUAGGC CGAA AGAGUCCU 2374
1025 AUGUGAGU U UGUUCUCC 186 GGAGAACA CUGAUGAG GCCGUUAGGC CGAA ACUCACAU 2375
1026 UGUGAGUU U GUUCUCCA 187 UGGAGAAC CUGAUGAG GCCGUUAGGC CGAA AACUCACA 2376
1029 GAGUUUGU U CUCCAAUC 188 GAUUGGAG CUGAUGAG GCCGUUAGGC CGAA ACAAACUC 2377
1030 AGUUUGUU C UCCAAUCC 189 GGAUUGGA CUGAUGAG GCCGUUAGGC CGAA AACAAACU 2378
1032 UUUGUUCU C CAAUCCCG 190 CGGGAUUG CUGAUGAG GCCGUUAGGC CGAA AGAACAAA 2379
1037 UCUCCAAU C CCGCCAGA 191 UCUGGCGG CUGAUGAG GCCGUUAGGC CGAA AUUGGAGA 2380
1057 AGAAGGCU U CUAUAAUG 192 CAUUAUAG CUGAUGAG GCCGUUAGGC CGAA AGCCUUCU 2381
1058 GAAGGCUU C UAUAAUGU 193 ACAUUAUA CUGAUGAG GCCGUUAGGC CGAA AAGCCUUC 2382
1060 AGGCUUCU A UAAUGUUU 194 AAACAUUA CUGAUGAG GCCGUUAGGC CGAA AGAAGCCU 2383
1062 GCUUCUAU A AUGUUUGC 195 GCAAACAU CUGAUGAG GCCGUUAGGC CGAA AUAGAAGC 2384
1067 UAUAAUGU U UGCACAAC 196 GUUGUGCA CUGAUGAG GCCGUUAGGC CGAA ACAUUAUA 2385
1068 AUAAUGUU U GCACAACA 197 UGUUGUGC CUGAUGAG GCCGUUAGGC CGAA AACAUUAU 2386
1080 CAACAUGU U GAUUCUAU 198 AUAGAAUC CUGAUGAG GCCGUUAGGC CGAA ACAUGUUG 2387
1084 AUGUUGAU U CUAUAGUU 199 AACUAUAG CUGAUGAG GCCGUUAGGC CGAA AUCAACAU 2388
1085 UGUUGAUU C UAUAGUUG 200 CAACUAUA CUGAUGAG GCCGUUAGGC CGAA AAUCAACA 2389
1087 UUGAUUCU A UAGUUGAA 201 UUCAACUA CUGAUGAG GCCGUUAGGC CGAA AGAAUCAA 2390
1089 GAUUCUAU A GUUGAAUU 202 AAUUCAAC CUGAUGAG GCCGUUAGGC CGAA AUAGAAUC 2391
1092 UCUAUAGU U GAAUUCUG 203 CAGAAUUC CUGAUGAG GCCGUUAGGC CGAA ACUAUAGA 2392
1097 AGUUGAAU U CUGUACAG 204 CUGUACAG CUGAUGAG GCCGUUAGGC CGAA AUUCAACU 2393
1098 GUUGAAUU C UGUACAGA 205 UCUGUACA CUGAUGAG GCCGUUAGGC CGAA AAUUCAAC 2394
1102 AAUUCUGU A CAGAACAA 206 UUGUUCUG CUGAUGAG GCCGUUAGGC CGAA ACAGAAUU 2395
1129 AAGAAGCU C CAAACAAG 207 CUUGUUUG CUGAUGAG GCCGUUAGGC CGAA AGCUUCUU 2396
1144 AGCAAAAU C AAAAAUGC 208 GCAUUUUU CUGAUGAG GCCGUUAGGC CGAA AUUUUGCU 2397
1156 AAUGCAAU C UCCGAAGC 209 GCUUCGGA CUGAUGAG GCCGUUAGGC CGAA AUUGCAUU 2398
1158 UGCAAUCU C CGAAGCAC 210 GUGCUUCG CUGAUGAG GCCGUUAGGC CGAA AGAUUGCA 2399
1179 GAAGUGAU c CGUGAUUC 211 GAAUCACG CUGAUGAG GCCGUUAGGC CGAA AUCACUUC 2400
1186 UCCGUGAU u CUGAGGAC 212 GUCCUCAG CUGAUGAG GCCGUUAGGC CGAA AUCACGGA 2401
1187 CCGUGAUU c UGAGGACU 213 AGUCCUCA CUGAUGAG GCCGUUAGGC CGAA AAUCACGG 2402
1196 UGAGGACU u UAAGAAAA 214 UUUUCUUA CUGAUGAG GCCGUUAGGC CGAA AGUCCUCA 2403
1197 GAGGACUU u AAGAAAAC 215 GUUUUCUU CUGAUGAG GCCGUUAGGC CGAA AAGUCCUC 2404
1198 AGGACUUU A AGAAAACC 216 GGUUUUCU CUGAUGAG GCCGUUAGGC CGAA AAAGUCCU 2405
1210 AAACCACU c CUAUGACA 217 UGUCAUAG CUGAUGAG GCCGUUAGGC CGAA AGUGGUUU 2406
1213 CCACUCCU A UGACAACA 218 UGUUGUCA CUGAUGAG GCCGUUAGGC CGAA AGGAGUGG 2407
1234 CACCAAAU C CCACCUUC 219 GAAGGUGG CUGAUGAG GCCGUUAGGC CGAA AUUUGGUG 2408
1241 UCCCACCU ϋ CUCAUUGC 220 GCAAUGAG CUGAUGAG GCCGUUAGGC CGAA AGGUGGGA 2409 1242 CCCACCUU C UCAUUGCU 221 AGCAAUGA CUGAUGAG GCCGUUAGGC CGAA AAGGUGGG 2410
1244 CACCUUCU C AUUGCUGC 222 GCAGCAAU CUGAUGAG GCCGUUAGGC CGAA AGAAGGUG 2411
1247 CUUCUCAU U GCUGCAGA 223 UCUGCAGC CUGAUGAG GCCGUUAGGC CGAA AUGAGAAG 2412
1257 CUGCAGAU U GGACAAAG 224 CUUUGUCC CUGAUGAG GCCGUUAGGC CGAA AUCUGCAG 2413
1269 CAAAGAAU U GUGUGUUU 225 AAACACAC CUGAUGAG GCCGUUAGGC CGAA AUUCUUUG 2414
1276 UUGUGUGU U UAGUCCUU 226 AAGGACUA CUGAUGAG GCCGUUAGGC CGAA ACACACAA 2415
1277 UGUGUGUU U AGUCCUUG 227 CAAGGACU CUGAUGAG GCCGUUAGGC CGAA AACACACA 2416
1278 GUGUGUUU A GUCCUUGA 228 UCAAGGAC CUGAUGAG GCCGUUAGGC CGAA AAACACAC 2417
1281 UGUUUAGU C CUUGACAA 229 UUGUCAAG CUGAUGAG GCCGUUAGGC CGAA ACUAAACA 2418
1284 UUAGUCCU U GACAAAUC 230 GAUUUGUC CUGAUGAG GCCGUUAGGC CGAA AGGACUAA 2419
1292 UGACAAAU C UGGAAGCA 231 UGCUUCCA CUGAUGAG GCCGUUAGGC CGAA AUUUGUCA 2420
1312 CGACUGGU A ACCGCCUC 232 GAGGCGGU CUGAUGAG GCCGUUAGGC CGAA ACCAGUCG 2421
1320 AACCGCCU C AAUCGACU 233 AGUCGAUU CUGAUGAG GCCGUUAGGC CGAA AGGCGGUU 2422
1324 GCCUCAAU C GACUGAAU 234 AUUCAGUC CUGAUGAG GCCGUUAGGC CGAA AUUGAGGC 2423
1333 GACUGAAU C AAGCAGGC 235 GCCUGCUU CUGAUGAG GCCGUUAGGC CGAA AUUCAGUC 2424
1347 GGCCAGCU U UUCCUGCU 236 AGCAGGAA. CUGAUGAG GCCGUUAGGC CGAA AGCUGGCC 2425
1348 GCCAGCUU U UCCUGCUG 237 CAGCAGGA CUGAUGAG GCCGUUAGGC CGAA AAGCUGGC 2426
1349 CCAGCUUU U CCUGCUGC 238 GCAGCAGG CUGAUGAG GCCGUUAGGC CGAA AAAGCUGG 2427
1350 CAGCUUUU C CUGCUGCA 239 UGCAGCAG CUGAUGAG GCCGUUAGGC CGAA AAAAGCUG 2428
1365 CAGACAGU U GAGCUGGG 240 CCCAGCUC CUGAUGAG GCCGUUAGGC CGAA ACUGUCUG 2429
1376 GCUGGGGU C CUGGGUUG 241 CAACCCAG CUGAUGAG GCCGUUAGGC CGAA ACCCCAGC 2430
1383 UCCUGGGU U GGGAUGGU 242 ACCAUCCC CUGAUGAG GCCGUUAGGC CGAA ACCCAGGA 2431
1397 GGUGACAU U UGACAGUG 243 CACUGUCA CUGAUGAG GCCGUUAGGC CGAA AUGUCACC 2432
1398 GUGACAUU U GACAGUGC 244 GCACUGUC CUGAUGAG GCCGUUAGGC CGAA AAUGUCAC 2433
1416 GCCCAUGU A CAAAGUGA 245 UCACUUUG CUGAUGAG GCCGUUAGGC CGAA ACAUGGGC 2434
1428 AGUGAACU C AUACAGAU 246 AUCUGUAU CUGAUGAG GCCGUUAGGC CGAA AGUUCACU 2435
1431 GAACUCAU A CAGAUAAA 247 UUUAUCUG CUGAUGAG GCCGUUAGGC CGAA AUGAGUUC 2436
1437 AUACAGAU A AACAGUGG 248 CCACUGUU CUGAUGAG GCCGUUAGGC CGAA AUCUGUAU 2437
1464 GACACACU C GCCAAAAG 249 CUUUUGGC CUGAUGAG GCCGUUAGGC CGAA AGUGUGUC 2438
1475 CAAAAGAU U ACCUGCAG 250 CUGCAGGU CUGAUGAG GCCGUUAGGC CGAA AUCUUUUG 2439
1476 AAAAGAUU A CCUGCAGC 251 GCUGCAGG CUGAUGAG GCCGUUAGGC CGAA AAUCUUUU 2440
1489 CAGCAGGU U CAGGAGGG 252 CCCUCCUG CUGAUGAG GCCGUUAGGC CGAA AGCUGCUG 2441
1490 AGCAGCUU C AGGAGGGA 253 UCCCUCCU CUGAUGAG GCCGUUAGGC CGAA AAGCUGCU 2442
1502 AGGGACGU C CAUCUGCA 254 UGCAGAUG CUGAUGAG GCCGUUAGGC CGAA ACGUCCCU 2443
1506 ACGUCCAU C UGCAGCGG 255 CCGCUGCA CUGAUGAG GCCGUUAGGC CGAA AUGGACGU 2444
1518 AGCGGGCU U CGAUCGGC 256 GCCGAUCG CUGAUGAG GCCGUUAGGC CGAA AGCCCGCU 2445
1519 GCGGGCUU C GAUCGGCA 257 UGCCGAUC CUGAUGAG GCCGUUAGGC CGAA AAGCCCGC 2446
1523 GCUUCGAU C GGCAUUUA 258 UAAAUGCC CUGAUGAG GCCGUUAGGC CGAA AUCGAAGC 2447
1529. AUCGGCAU. U UACUGUGA 259 -UCACAGUA. CUGAUGAG GCCGUUAGGC CGAA AUGCCGAU 2448
1530 UCGGCAUU U ACUGUGAU 260 AUCACAGU CUGAUGAG GCCGUUAGGC CGAA AAUGCCGA 2449
1531 CGGCAUUU A CUGUGAUU 261 AAUCACAG CUGAUGAG GCCGUUAGGC CGAA AAAUGCCG 2450
1539 ACUGUGAU U AGGAAGAA 262 UUCUUCCU CUGAUGAG GCCGUUAGGC CGAA AUCACAGU 2451
1540 CUGUGAUU A GGAAGAAA 263 UUUCUUCC CUGAUGAG GCCGUUAGGC CGAA AAUCACAG 2452
1550 GAAGAAAU A UCCAACUG 264 CAGUUGGA CUGAUGAG GCCGUUAGGC CGAA AUUUCUUC 2453
1552 AGAAAUAU C CAACUGAU 265 AUCAGUUG CUGAUGAG GCCGUUAGGC CGAA AUAUUUCU 2454
1565 UGAUGGAU C UGAAAUUG 266 CAAUUUCA CUGAUGAG GCCGUUAGGC CGAA AUCCAUCA 2455
1572 UCUGAAAU U GUGCUGCU 267 AGCAGCAC CUGAUGAG GCCGUUAGGC CGAA AUUUCAGA 2456
1603 ACAACACU A UAAGUGGG 268 CCCACUUA CUGAUGAG GCCGUUAGGC CGAA AGUGUUGU 2457
1605 AACACUAU A AGUGGGUG 269 CACCCACU CUGAUGAG GCCGUUAGGC CGAA AUAGUGUU 2458
1616 UGGGUGCU U UAACGAGG 270 CCUCGUUA CUGAUGAG GCCGUUAGGC CGAA AGCACCCA 2459
1617 GGGUGCUU U AACGAGGU 271 ACCUCGUU CUGAUGAG GCCGUUAGGC CGAA AAGCACCC 2460
1618 GGUGCUUU A ACGAGGUC 272 GACCUCGU CUGAUGAG GCCGUUAGGC CGAA AAAGCACC 2461
1626 AACGAGGU C AAACAAAG 273 CUUUGUUU CUGAUGAG GCCGUUAGGC CGAA ACCUCGUU 2462
1644 GGUGCCAU C AUCCACAC 274 GUGUGGAU CUGAUGAG GCCGUUAGGC CGAA AUGGCACC 2463
1647 GCCAUCAU C CACACAGU 275 ACUGUGUG CUGAUGAG GCCGUUAGGC CGAA AUGAUGGC 2464
1656 CACACAGU C GCUUUGGG 276 CCCAAAGC CUGAUGAG GCCGUUAGGC CGAA ACUGUGUG 2465 1660 CAGUCGCU U UGGGGCCC 277 GGGCCCCA CUGAUGAG GCCGUUAGGC CGAA. AGCGACUG 2466
1661 AGUCGCUU U GGGGCCCU 278 AGGGCCCC CUGAUGAG GCCGUUAGGC CGAA AAGCGACU 2467
1670 GGGGCCCU C UGCAGCUC 279 GAGCUGCA CUGAUGAG GCCGUUAGGC CGAA AGGGCCCC 2468
1678 CUGCAGCU C AAGAACUA 280 UAGUUCUU CUGAUGAG GCCGUUAGGC CGAA AGCUGCAG 2469
1686 CAAGAACU A GAGGAGCU 281 AGCUCCUC CUGAUGAG GCCGUUAGGC CGAA AGUUCUUG 2470
1697 GGAGCUGU C CAAAAUGA 282 UCAUUUUG CUGAUGAG GCCGUUAGGC CGAA ACAGCUCC 2471
1714 CAGGAGGU u UACAGACA 283 UGUCUGUA CUGAUGAG GCCGUUAGGC CGAA ACCUCCUG 2472
1715 AGGAGGUU u ACAGACAU 284 AUGUCUGU CUGAUGAG GCCGUUAGGC CGAA AACCUCCU 2473
1716 GGAGGUUU A CAGACAUA 285 UAUGUCUG CUGAUGAG GCCGUUAGGC CGAA AAACCUCC 2474
1724 ACAGACAU A UGCUUCAG 286 CUGAAGCA CUGAUGAG GCCGUUAGGC CGAA AUGUCUGU 2475
1729 CAUAUGCU U CAGAUCAA 287 UUGAUCUG CUGAUGAG GCCGUUAGGC CGAA AGCAUAUG 2476
1730 AUAUGCUU C AGAUCAAG 288 CUUGAUCU CUGAUGAG GCCGUUAGGC CGAA AAGCAUAU 2477
1735 CUUCAGAU C AAGUUCAG 289 CUGAACUU CUGAUGAG GCCGUUAGGC CGAA AUCUGAAG 2478
1740 GAUCAAGU u CAGAACAA 290^ UUGUUCUG CUGAUGAG GCCGUUAGGC CGAA ACUUGAUC 2479
1741 AUCAAGUU c AGAACAAU 291 AUUGUUCU CUGAUGAG GCCGUUAGGC CGAA AACUUGAU 2480
1755 AAUGGCCU c AUUGAUGC 292 GCAUCAAU CUGAUGAG GCCGUUAGGC CGAA AGGCCAUU 2481
1758 GGCCUCAU u GAUGCUUU 293 AAAGCAUC CUGAUGAG GCCGUUAGGC CGAA AUGAGGCC 2482
1765 UUGAUGCU u UUGGGGCC 294 GGCCCCAA CUGAUGAG GCCGUUAGGC CGAA AGCAUCAA 2483
1766 UGAUGCUU u UGGGGCCC 295 GGGCCCCA CUGAUGAG GCCGUUAGGC CGAA AAGCAUCA 2484
1767 GAUGCUUU u GGGGCCCU 296 AGGGCCCC CUGAUGAG GCCGUUAGGC CGAA AAAGCAUC 2485
1776 GGGGCCCU u UCAUCAGG 297 CCUGAUGA CUGAUGAG GCCGUUAGGC CGAA AGGGCCCC 2486
1777 GGGCCCUU u CAUCAGGA 298 UCCUGAUG CUGAUGAG GCCGUUAGGC CGAA AAGGGCCC 2487
1778 GGCCCUUU c AUCAGGAA 299 UUCCUGAU CUGAUGAG GCCGUUAGGC CGAA AAAGGGCC 2488
1781 CCUUUCAU c AGGAAAUG 300 CAUUUCCU CUGAUGAG GCCGUUAGGC CGAA AUGAAAGG 2489
1797 GGAGCUGU c UCUCAGCG 301 CGCUGAGA CUGAUGAG GCCGUUAGGC CGAA ACAGCUCC 2490
1799 AGCUGUCU c UCAGCGCU 302 AGCGCUGA CUGAUGAG GCCGUUAGGC CGAA AGACAGCU 2491
1801 CUGUCUCU c AGCGCUCC 303 GGAGCGCU CUGAUGAG GCCGUUAGGC CGAA AGAGACAG 2492
1808 UCAGCGCU c CAUCCAGC 304 GCUGGAUG CUGAUGAG GCCGUUAGGC CGAA AGCGCUGA 2493
1812 CGCUCCAU c CAGCUUGA 305 UCAAGCUG CUGAUGAG GCCGUUAGGC CGAA AUGGAGCG 2494
1818 AUCCAGCU u GAGAGUAA 306 UUACUCUC CUGAUGAG GCCGUUAGGC CGAA AGCUGGAU 2495
1825 UUGAGAGU A AGGGAUUA 307 UAAUCCCU CUGAUGAG GCCGUUAGGC CGAA ACUCUCAA 2496
1832 UAAGGGAU u AACCCUCC 308 GGAGGGUU CUGAUGAG GCCGUUAGGC CGAA AUCCCUUA 2497
1833 AAGGGAUU A ACCCUCCA 309 UGGAGGGU CUGAUGAG GCCGUUAGGC CGAA AAUCCCUU 2498
1839 UUAACCCU c CAGAACAG 310 CUGUUCUG CUGAUGAG GCCGUUAGGC CGAA AGGGUUAA 2499
1872 ACAGUGAU c GUGGACAG 311 CUGUCCAC CUGAUGAG GCCGUUAGGC CGAA AUCACUGU 2500
1900 AGGACACU u UGUUUCUU 312 AAGAAACA CUGAUGAG GCCGUUAGGC CGAA AGUGUCCU 2501
1901 GGACACUU u GUUUCUUA 313 UAAGAAAC CUGAUGAG GCCGUUAGGC CGAA AAGUGUCC 2502
1904 CACUUUGU u UCUUAUCA 314 UGAUAAGA CUGAUGAG GCCGUUAGGC CGAA ACAAAGUG 2503
1905 ACUUUGUU u CUUAUCAC 315 GUGAUAAG CUGAUGAG GCCGUUAGGC CGAA AACAAAGU 2504
1906 CUUUGUUU c UUAUCACC 316 GGUGAUAA CUGAUGAG GCCGUUAGGC CGAA AAACAAAG 2505
1908 UUGUUUCU u AUCACCUG 317 CAGGUGAU CUGAUGAG GCCGUUAGGC CGAA AGAAACAA 2506
1909 UGUUUCUU A UCACCUGG 318 CCAGGUGA CUGAUGAG GCCGUUAGGC CGAA AAGAAACA 2507
1911 UUUCUUAU c ACCUGGAC 319 GUCCAGGU CUGAUGAG GCCGUUAGGC CGAA AUAAGAAA 2508
1930 CGCAGCCU c CCCAAAUC 320 GAUUUGGG CUGAUGAG GCCGUUAGGC CGAA AGGCUGCG 2509
1938 CCCCAAAU c CUUCUCUG 321 CAGAGAAG CUGAUGAG GCCGUUAGGC CGAA AUUUGGGG 2510
1941 CAAAUCCU u CUCUGGGA 322 UCCCAGAG CUGAUGAG GCCGUUAGGC CGAA AGGAUUUG 2511
1942 AAAUCCUU c UCUGGGAU 323 AUCCCAGA CUGAUGAG GCCGUUAGGC CGAA AAGGAUUU 2512
1944 AUCCUUCU c UGGGAUCC 324 GGAUCCCA CUGAUGAG GCCGUUAGGC CGAA AGAAGGAU 2513
1951 UCUGGGAU c CCAGUGGA 325 UCCACUGG CUGAUGAG GCCGUUAGGC CGAA AUCCCAGA 2514
1976 AGGUGGCU u UGUAGUGG 326 CCACUACA CUGAUGAG GCCGUUAGGC CGAA AGCCACCU 2515
1977 GGUGGCUU u GUAGUGGA 327 UCCACUAC CUGAUGAG GCCGUUAGGC CGAA AAGCCACC 2516
1980 GGCUUUGU A GUGGACAA 328 UUGUCCAC CUGAUGAG GCCGUUAGGC CGAA ACAAAGCC 2517
2006 AAUGGCCU A CCUCCAAA 329 UUUGGAGG CUGAUGAG GCCGUUAGGC CGAA AGGCCAUU 2518
2010 GCCUACCU c CAAAUCCC 330 GGGAUUUG CUGAUGAG GCCGUUAGGC CGAA AGGUAGGC 2519
2016 CUCCAAAU C CCAGGCAU 331 AUGCCUGG CUGAUGAG GCCGUUAGGC CGAA AUUUGGAG 2520
2025 CCAGGCAU u GCUAAGGU 332 ACCUUAGC CUGAUGAG GCCGUUAGGC CGAA AUGCCUGG 2521 2029 GCAUUGCU A AGGUUGGC 333 GCCAACCU CUGAUGAG GCCGUUAGGC CGAA AGCAAUGC 2522
2034 GCUAAGGU U GGCACUUG 334 CAAGUGCC CUGAUGAG GCCGUUAGGC CGAA ACCUUAGC 2523
2041 UUGGCACU U GGAAAUAC 335 GUAUUUCC CUGAUGAG GCCGUUAGGC CGAA AGUGCCAA 2524
2048 UUGGAAAU A CAGUCUGC 336 GCAGACUG CUGAUGAG GCCGUUAGGC CGAA AϋϋUCCAA 2525
2053 AAUACAGU C UGCAAGCA 337 UGCUUGCA CUGAUGAG GCCGUUAGGC CGAA ACUGUAUU 2526
2066 AGCAAGCU C ACAAACCU 338 AGGUUUGU CUGAUGAG GCCGUUAGGC CGAA AGCUUGCU 2527
2075 ACAAACCU U GACCCUGA 339 UCAGGGUC CUGAUGAG GCCGUUAGGC CGAA AGGUUUGU 2528
2088 CUGACUGU C ACGUCCCG 340 CGGGACGU CUGAUGAG GCCGUUAGGC CGAA ACAGUCAG 2529
2093 UGUCACGU C CCGUGCGU 341 ACGCACGG CUGAUGAG GCCGUUAGGC CGAA ACGUGACA 2530
2102 CCGUGCGU C CAAUGCUA 342 UAGCAUUG CUGAUGAG GCCGUUAGGC CGAA ACGCACGG 2531
2110 CCAAUGCU A CCCUGCCU 343 AGGCAGGG CUGAUGAG GCCGUUAGGC CGAA AGCAUUGG 2532
2119 CCCUGCCU C CAAUUACA 344 UGUAAUUG CUGAUGAG GCCGUUAGGC CGAA AGGCAGGG 2533
2124 CCUCCAAU U ACAGUGAC 345 GUCACUGU CUGAUGAG GCCGUUAGGC CGAA AUUGGAGG 2534
2125 CUCCAAUU A CAGUGACU 346 AGUCACUG CUGAUGAG GCCGUUAGGC CGAA AAUUGGAG 2535
2134 CAGUGACU U CCAAAACG 347 cσuuuuGG CUGAUGAG GCCGUUAGGC CGAA AGUCACUG 2536
2135 AGUGACUU C CAAAACGA 348 ucGuuuuσ CUGAUGAG GCCGUUAGGC CGAA AAGUCACϋ 2537
2162 CAGCAAAU U CCCCAGCC 349 GGCUGGGG CUGAUGAG GCCGUUAGGC CGAA AUUUGCUG 2538
2163 AGCAAAUU C CCCAGCCC 350 GGGCUGGG CUGAUGAG GCCGUUAGGC CGAA AAUUUGCU 2539
2173 CCAGCCCU C UGGUAGUU 351 AACUACCA CUGAUGAG GCCGUUAGGC CGAA AGGGCUGG 2540
2178 CCUCUGGU A GUUUAUGC 352 GCAUAAAC CUGAUGAG GCCGUUAGGC CGAA ACCAGAGG 2541
2181 CUGGUAGU U UAUGCAAA 353 UUUGCAUA CUGAUGAG GCCGUUAGGC CGAA ACUACCAG 2542
2182 UGGUAGUU U AUGCAAAU 354 AUUUGCAU CUGAUGAG GCCGUUAGGC CGAA AACUACCA 2543
2183 GGUAGUUU A UGCAAAUA 355 UAUUUGCA CUGAUGAG GCCGUUAGGC CGAA AAACUACC 2544
2191 AUGCAAAU A UUCGCCAA 356 UϋGGCGAA CUGAUGAG GCCGUUAGGC CGAA AUUUGCAU 2545
2193 GCAAAUAU U CGCCAAGG 357 CCUUGGCG CUGAUGAG GCCGUUAGGC CGAA AUAUUUGC 2546
2194 CAAAUAUU C GCCAAGGA 358 UCCUUGGC CUGAUGAG GCCGUUAGGC CGAA AAUAUUUG 2547
2207 AGGAGCCU C CCCAAUUC 359 GAAUUGGG CUGAUGAG GCCGUUAGGC CGAA AGGCUCCU 2548
2214 UCCCCAAU U CUCAGGGC 360 GCCCUGAG CUGAUGAG GCCGUUAGGC CGAA AUUGGGGA 2549
2215 CCCCAAUU c UCAGGGCC 361 GGCCCUGA CUGAUGAG GCCGUUAGGC CGAA AAUUGGGG 2550
2217 CCAAUUCU c AGGGCCAG 362 CUGGCCCU CUGAUGAG GCCGUUAGGC CGAA AGAAUUGG 2551
2229 GCCAGUGU c ACAGCCCU 363 AGGGCUGU CUGAUGAG GCCGUUAGGC CGAA ACACUGGC 2552
2241 GCCCUGAU u GAAUCAGU 364 ACUGAUUC CUGAUGAG GCCGUUAGGC CGAA AUCAGGGC 2553
2246 GAUUGAAU c AGUGAAUG 365 CAUUCACU CUGAUGAG GCCGUUAGGC CGAA AUUCAAUC 2554
2265 AAAACAGU u ACCUUGGA 366 UCCAAGGU CUGAUGAG GCCGUUAGGC CGAA ACUGUUUU 2555
2266 AAACAGUU A CCUUGGAA 367 UUCCAAGG CUGAUGAG GCCGUUAGGC CGAA AACUGUUU 2556
2270 AGUUACCU u GGAACUAC 368 GUAGUUCC CUGAUGAG GCCGUUAGGC CGAA AGGUAACU 2557
2277 UUGGAACU A CUGGAUAA 369 UUAUCCAG CUGAUGAG GCCGUUAGGC CGAA AGUUCCAA 2558
2284 UACUGGAU A AUGGAGCA 370 UGCUCCAU CUGAUGAG GCCGUUAGGC CGAA AUCCAGUA 2559
2305 CUGAUGCU A .CUAAGGAU 371 AUCCUUAG .CUGAUGAG G.C.CGUUAGGC CGAA AGCAUCAG 2560
2308 AUGCUACU A AGGAUGAC 372 GUCAUCCU CUGAUGAG GCCGUUAGGC CGAA AGUAGCAU 2561
2322 GACGGUGU C UACUCAAG 373 CUUGAGUA CUGAUGAG GCCGUUAGGC CGAA ACACCGUC 2562
2324 CGGUGUCU A CUCAAGGU 374 ACCUUGAG CUGAUGAG GCCGUUAGGC CGAA AGACACCG 2563
2327 UGUCUACU C AAGGUAUU 375 AAUACCUU CUGAUGAG GCCGUUAGGC CGAA AGUAGACA 2564
2333 CUCAAGGU A UUUCACAA 376 UUGUGAAA CUGAUGAG GCCGUUAGGC CGAA ACCUUGAG 2565
2335 CAAGGUAU U UCACAACU 377 AGUUGUGA CUGAUGAG GCCGUUAGGC CGAA AUACCUUG 2566
2336 AAGGUAUU U CACAACUU 378 AAGUUGUG CUGAUGAG GCCGUUAGGC CGAA AAUACCUU 2567
2337 AGGUAUUU C ACAACUUA 379 UAAGUUGU CUGAUGAG GCCGUUAGGC CGAA AAAUACCU 2568
2344 UCACAACU u AUGACACG 380 CGUGUCAU CUGAUGAG GCCGUUAGGC CGAA AGUUGUGA 2569
2345 CACAACUU A UGACACGA 381 UCGUGUCA CUGAUGAG GCCGUUAGGC CGAA AAGUUGUG 2570
2359 CGAAUGGU A GAUACAGU 382 ACUGUAUC CUGAUGAG GCCGUUAGGC CGAA ACCAUUCG 2571
2363 UGGUAGAU A .CAGUGUAA 383 UUACACUG CUGAUGAG GCCGUUAGGC CGAA AUCUACCA 2572
2370 UACAGUGU A AAAGUGCG 384 CGCACUUU CUGAUGAG GCCGUUAGGC CGAA ACACUGUA 2573
2383 UGCGGGCU C UGGGAGGA 385 UCCUCCCA CUGAUGAG GCCGUUAGGC CGAA AGCCCGCA 2574
2394 GGAGGAGU U AACGCAGC 386 GCUGCGUU CUGAUGAG GCCGUUAGGC CGAA ACUCCUCC 2575
2395 GAGGAGUU A ACGCAGCC 387 GGCUGCGU CUGAUGAG GCCGUUAGGC CGAA AACUCCUC 2576
2418 AGAGUGAU A CCCCAGCA 388 UGCUGGGG CUGAUGAG GCCGUUAGGC CGAA AUCACUCU 2577 2441 AGCACUGU A CAUACCUG 389 CAGGUAUG CUGAUGAG GCCGUUAGGC CGAA ACAGUGCU 2578
2445 CUGUACAU A CCUGGCUG 390 CAGCCAGG CUGAUGAG GCCGUUAGGC CGAA AUGUACAG 2579
2457 GGCUGGAU U GAGAAUGA 391 UCAUUCUC CUGAUGAG GCCGUUAGGC CGAA AUCCAGCC 2580
2472 GAUGAAAU A CAAUGGAA 392 UUCCAUUG CUGAUGAG GCCGUUAGGC CGAA AUUUCAUC 2581
2482 AAUGGAAU C CACCAAGA 393 UCUUGGUG CUGAUGAG GCCGUUAGGC CGAA AUUCCAUU 2582
2499 CCUGAAAU U AAUAAGGA 394 UCCUUAUU CUGAUGAG GCCGUUAGGC CGAA AUUUCAGG 2583
2500 CUGAAAUU A AUAAGGAU 395 AUCCUUAU CUGAUGAG GCCGUUAGGC CGAA AAUUUCAG 2584
2503 AAAUUAAU A AGGAUGAU 396 AUCAUCCU CUGAUGAG GCCGUUAGGC CGAA AUUAAUUU 2585
2514 GAUGAUGU U CAACACAA 397 UUGUGUUG CUGAUGAG GCCGUUAGGC CGAA ACAUCAUC 2586
2515 AUGAUGUU C AACACAAG 398 CUUGUGUU CUGAUGAG GCCGUUAGGC CGAA AACAUCAU 2587
2533 AAGUGUGU U UCAGCAGA 399 UCUGCUGA CUGAUGAG GCCGUUAGGC CGAA ACACACUU 2588
2534 AGUGUGUU U CAGCAGAA 400 UUCUGCUG CUGAUGAG GCCGUUAGGC CGAA AACACACU 2589
2535 GUGUGUUU C AGCAGAAC 401 GUUCUGCU CUGAUGAG GCCGUUAGGC CGAA AAACACAC 2590
2546 CAGAACAU C CUCGGGAG 402 CUCCCGAG CUGAUGAG GCCGUUAGGC CGAA AUGUUCUG 2591
2549 AACAUCCU C GGGAGGCU 403 AGCCUCCC CUGAUGAG GCCGUUAGGC CGAA AGGAUGUU 2592
2558 GGGAGGCU C- AUUUGUGG -404 CCACAAAU CUGAUGAG--GCCGUUAGGC CGAA AGCCUCCC 2593
2561 AGGCUCAU U UGUGGCUU 405 AAGCCACA CUGAUGAG GCCGUUAGGC CGAA AUGAGCCU 2594
2562 GGCUCAUU U GUGGCUUC 406 GAAGCCAC CUGAUGAG GCCGUUAGGC CGAA AAUGAGCC 2595
2569 UUGUGGCU U CUGAUGUC 407 GACAUCAG CUGAUGAG GCCGUUAGGC CGAA AGCCACAA 2596
2570 UGUGGCUU C UGAUGUCC 408 GGACAUCA CUGAUGAG GCCGUUAGGC CGAA AAGCCACA 2597
2577 UCUGAUGU C CCAAAUGC 409 GCAUUUGG CUGAUGAG GCCGUUAGGC CGAA ACAUCAGA 2598
2587 CAAAUGCU C CCAUACCU 410 AGGUAUGG CUGAUGAG GCCGUUAGGC CGAA AGCAUUUG 2599
2592 GCUCCCAU A CCUGAUCU 411 AGAUCAGG CUGAUGAG GCCGUUAGGC CGAA AUGGGAGC 2600
2599 UACCUGAU C UCUUCCCA 412 UGGGAAGA CUGAUGAG GCCGUUAGGC CGAA AUCAGGUA 2601
2601 CCUGAUCU C UUCCCACC 413 GGUGGGAA CUGAUGAG GCCGUUAGGC CGAA AGAUCAGG 2602
2603 UGAUCUCU U CCCACCUG 414 CAGGUGGG CUGAUGAG GCCGUUAGGC CGAA AGAGAUCA 2603
2604 GAUCUCUU C CCACCUGG 415 CCAGGUGG CUGAUGAG GCCGUUAGGC CGAA AAGAGAUC 2604
2619 GGCCAAAU C ACCGACCU 416 AGGUCGGU CUGAUGAG GCCGUUAGGC CGAA AUUUGGCC 2605
2640 GCGGAAAU U CACGGGGG 417 CCCCCGUG CUGAUGAG GCCGUUAGGC CGAA AUUUCCGC 2606
2641 CGGAAAUU C ACGGGGGC 418 GCCCCCGU CUGAUGAG GCCGUUAGGC CGAA AAUUUCCG 2607
2653 GGGGCAGU C UCAUUAAU 419 AUUAAUGA CUGAUGAG GCCGUUAGGC CGAA ACUGCCCC 2608
2655 GGCAGUCU C AUUAAUCU 420 AGAUUAAU CUGAUGAG GCCGUUAGGC CGAA AGACUGCC 2609
2658 AGUCUCAU U AAUCUGAC 421 GUCAGAUU CUGAUGAG GCCGUUAGGC CGAA AUGAGACU 2610
2659 GUCUCAUU A AUCUGACU 422 AGUCAGAU CUGAUGAG GCCGUUAGGC CGAA AAUGAGAC 2611
2662 UCAUUAAU C UGACUUGG 423 CCAAGUCA CUGAUGAG GCCGUUAGGC CGAA AUUAAUGA 2612
2668 AUCUGACU U GGACAGCU 424 AGCUGUCC CUGAUGAG GCCGUUAGGC CGAA AGUCAGAU 2613
2677 GGACAGCU C CUGGGGAU 425 AUCCCCAG CUGAUGAG GCCGUUAGGC CGAA AGCUGUCC 2614
2689 GGGAUGAU U AUGACCAU 426 AUGGUCAU CUGAUGAG GCCGUUAGGC CGAA AUCAUCCC 2615
2690 GGAUGAUU A UGACCAUG 427 _ CAUGGUCA. -CUGAUGAG GCCGUUAGGC .CGAA-AAUCAUCC 2616
2707 GAACAGCU C ACAAGUAU 428 AUACUUGU CUGAUGAG GCCGUUAGGC CGAA AGCUGUUC 2617
2714 UCACAAGU A UAUCAUUC 429 GAAUGAUA CUGAUGAG GCCGUUAGGC CGAA ACUUGUGA 2618
2716 ACAAGUAU A UCAUUCGA 430 UCGAAUGA CUGAUGAG GCCGUUAGGC CGAA AUACUUGU 2619
2718 AAGUAUAU C AUUCGAAU 431 AUUCGAAU CUGAUGAG GCCGUUAGGC CGAA AUAUACUU 2620
2721 UAUAUCAU U CGAAUAAG 432 CUUAUUCG CUGAUGAG GCCGUUAGGC CGAA AUGAUAUA 2621
2722 AUAUCAUU C GAAUAAGU 433 ACUUAUUC CUGAUGAG GCCGUUAGGC CGAA AAUGAUAU 2622
2727 AUUCGAAU A AGUACAAG 434 CUUGUACU CUGAUGAG GCCGUUAGGC CGAA AUUCGAAU 2623
2731 GAAUAAGU A CAAGUAUU 435 AAUACUUG CUGAUGAG GCCGUUAGGC CGAA ACUUAUUC 2624
2737 GUACAAGU A UUCUUGAU 436 AUCAAGAA CUGAUGAG GCCGUUAGGC CGAA ACUUGUAC 2625
2739 ACAAGUAU U CUUGAUCU 437 AGAUCAAG CUGAUGAG GCCGUUAGGC CGAA AUACUUGU 2626
2740 CAAGUAUU C UUGAUCUC 438 GAGAUCAA CUGAUGAG GCCGUUAGGC CGAA AAUACUUG 2627
2742 AGUAUUCU U GAUCUCAG 439 CUGAGAUC CUGAUGAG GCCGUUAGGC CGAA AGAAUACU 2628
2746 UUCUUGAU C UCAGAGAC 440 GUCUCUGA CUGAUGAG GCCGUUAGGC CGAA AUCAAGAA 2629
2748 CUUGAUCU C AGAGACAA 441 UUGUCUCU CUGAUGAG GCCGUUAGGC CGAA AGAUCAAG 2630
2759 AGACAAGU U CAAUGAAU 442 AUUCAUUG CUGAUGAG GCCGUUAGGC CGAA ACUUGUCU 2631
2760 GACAAGUU C AAUGAAUC 443 GAUUCAUU CUGAUGAG GCCGUUAGGC CGAA AACUUGUC 2632
2768 CAAUGAAU C UCUUCAAG 444 CUUGAAGA CUGAUGAG GCCGUUAGGC CGAA AUUCAUUG 2633 2770 AUGAAUCU C UUCAAGUG 445 CACUUGAA CUGAUGAG GCCGUUAGGC CGAA AGAUUCAU 2634
2772 GAAUCUCU U CAAGUGAA 446 UUCACUUG CUGAUGAG GCCGUUAGGC CGAA AGAGAUUC 2635
2773 AAUCUCUU C AAGUGAAU 447 AUUCACUU CUGAUGAG GCCGUUAGGC CGAA AAGAGAUU 2636
2782 AAGUGAAU A CUACUGCU 448 AGCAGUAG CUGAUGAG GCCGUUAGGC CGAA AUUCACUU 2637
2785 UGAAUACU A CUGCUCUC 449 GAGAGCAG CUGAUGAG GCCGUUAGGC CGAA AGUAUUCA 2638
2791 CUACUGCU C UCAUCCCA 450 UGGGAUGA CUGAUGAG GCCGUUAGGC CGAA AGCAGUAG 2639
2793 ACUGCUCU C AUCCCAAA 451 UUUGGGAU CUGAUGAG GCCGUUAGGC CGAA AGAGCAGU 2640
2796 GCUCUCAU C CCAAAGGA 452 UCCUUUGG CUGAUGAG GCCGUUAGGC CGAA AUGAGAGC 2641
2813 AGCCAACU C UGAGGAAG 453 CUUCCUCA CUGAUGAG GCCGUUAGGC CGAA AGUUGGCU 2642
2823 GAGGAAGU C UUUUUGUU 454 AACAAAAA CUGAUGAG GCCGUUAGGC CGAA ACUUCCUC 2643
2825 GGAAGUCU U UUUGUUUA 455 UAAACAAA CUGAUGAG GCCGUUAGGC CGAA AGACUUCC 2644
2826 GAAGUCUU U UUGUUUAA 456 UUAAACAA CUGAUGAG GCCGUUAGGC CGAA AAGACUUC 2645
2827 AAGUCUUU U UGUUUAAA 457 UUUAAACA CUGAUGAG GCCGUUAGGC CGAA AAAGACUU 2646
2828 AGUCUUUU U GUUUAAAC 458 GUUUAAAC CUGAUGAG GCCGUUAGGC CGAA AAAAGACU 2647
2831 CUUUUUGU u UAAACCAG 459 CUGGUUUA CUGAUGAG GCCGUUAGGC CGAA ACAAAAAG 2648
2832 UUUUUGUU u AAACCAGA -460 UCUGGUUU CUGAUGAG "GCCGUUAGGC CGAA AACAAAAA 2649
2833 UUUUGUUU A AACCAGAA 461 UUCUGGUU CUGAUGAG GCCGUUAGGC CGAA AAACAAAA 2650
2847 GAAAACAU U ACUUUUGA 462 UCAAAAGU CUGAUGAG GCCGUUAGGC CGAA AUGUUUUC 2651
2848 AAAACAUU A CUUUUGAA 463 UUCAAAAG CUGAUGAG GCCGUUAGGC CGAA AAUGUUUU 2652
2851 ACAUUACU U UUGAAAAU 464 AUUUUCAA CUGAUGAG GCCGUUAGGC CGAA AGUAAUGU 2653
2852 CAUUACUU U UGAAAAUG 465 CAUUUUCA CUGAUGAG GCCGUUAGGC CGAA AAGUAAUG 2654
2853 AUUACUUU U GAAAAUGG 466 CCAUUUUC CUGAUGAG GCCGUUAGGC CGAA AAAGUAAU 2655
2869 GCACAGAU C UUUUCAUU 467 AAUGAAAA CUGAUGAG GCCGUUAGGC CGAA AUCUGUGC 2656
2871 ACAGAUCU U UUCAUUGC 468 GCAAUGAA CUGAUGAG GCCGUUAGGC CGAA AGAUCUGU 2657
2872 CAGAUCUU U UCAUUGCU 469 AGCAAUGA CUGAUGAG GCCGUUAGGC CGAA AAGAUCUG 2658
2873 AGAUCUUU U CAUUGCUA 470 UAGCAAUG CUGAUGAG GCCGUUAGGC CGAA AAAGAUCU 2659
2874 GAUCUUUU C AUUGCUAU 471 AUAGCAAU CUGAUGAG GCCGUUAGGC CGAA AAAAGAUC 2660
2877 CUUUUCAU u GCUAUUCA 472 UGAAUAGC CUGAUGAG GCCGUUAGGC CGAA AUGAAAAG 2661
2881 UCAUUGCU A UUCAGGCU 473 AGCCUGAA CUGAUGAG GCCGUUAGGC CGAA AGCAAUGA 2662
2883 AUUGCUAU U CAGGCUGU 474 ACAGCCUG CUGAUGAG GCCGUUAGGC CGAA AUAGCAAU 2663
2884 UUGCUAUU C AGGCUGUU 475 AACAGCCU CUGAUGAG GCCGUUAGGC CGAA AAUAGCAA 2664
2892 CAGGCUGU U GAUAAGGU 476 ACCUUAUC CUGAUGAG GCCGUUAGGC CGAA ACAGCCUG 2665
2896 CUGUUGAU A AGGUCGAU 477 AUCGACCU CUGAUGAG GCCGUUAGGC CGAA AUCAACAG 2666
2901 GAUAAGGU c GAUCUGAA 478 UUCAGAUC CUGAUGAG GCCGUUAGGC CGAA ACCUUAUC 2667
2905 AGGUCGAU c UGAAAUCA 479 UGAUUUCA CUGAUGAG GCCGUUAGGC CGAA AUCGACCU 2668
2912 UCUGAAAU c AGAAAUAU 480 AUAUUUCU CUGAUGAG GCCGUUAGGC CGAA AUUUCAGA 2669
2919 UCAGAAAU A UCCAACAU 481 AUGUUGGA CUGAUGAG GCCGUUAGGC CGAA AUUUCUGA 2670
2921 AGAAAUAU C CAACAUUG 482 CAAUGUUG CUGAUGAG GCCGUUAGGC CGAA AUAUUUCU 2671
2928 UCCAACAU U. GCACGAGU .483 ACUCGUGC -CUGAUGAG ..GCCGUUAGGC -CGAA -AUGUUGGA 2672
2937 GCACGAGU A UCUUUGUU 484 AACAAAGA CUGAUGAG GCCGUUAGGC CGAA ACUCGUGC 2673
2939 ACGAGUAU c UUUGUUUA 485 UAAACAAA CUGAUGAG GCCGUUAGGC CGAA AUACUCGU 2674
2941 GAGUAUCU U UGUUUAUU 486 AAUAAACA CUGAUGAG GCCGUUAGGC CGAA AGAUACUC 2675
2942 AGUAUCUU u GUUUAUUC 487 GAAUAAAC CUGAUGAG GCCGUUAGGC CGAA AAGAUACU 2676
2945 AUCUUUGU u UAUUCCUC 488 GAGGAAUA CUGAUGAG GCCGUUAGGC CGAA ACAAAGAU 2677
2946 UCUUUGUU u AUUCCUCC 489 GGAGGAAU CUGAUGAG GCCGUUAGGC CGAA AACAAAGA 2678
2947 CUUUGUUU A UUCCUCCA 490 UGGAGGAA CUGAUGAG GCCGUUAGGC CGAA AAACAAAG 2679
2949 UUGUUUAU u CCUCCACA 491 UGUGGAGG CUGAUGAG GCCGUUAGGC CGAA AUAAACAA 2680
2950 UGUUUAUU C CUCCACAG 492 CUGUGGAG CUGAUGAG GCCGUUAGGC CGAA AAUAAACA 2681
2953 UUAUUCCU C CACAGACU 493 AGUCUGUG CUGAUGAG GCCGUUAGGC CGAA AGGAAUAA 2682
2962 CACAGACU c CGCCAGAG 494 CUCUGGCG CUGAUGAG GCCGUUAGGC CGAA AGUCUGUG 2683
2977 AGACACCU A GUCCUGAU 495 AUCAGGAC CUGAUGAG GCCGUUAGGC CGAA AGGUGUCU 2684
2980 CACCUAGU c CUGAUGAA 496 UUCAUCAG CUGAUGAG GCCGUUAGGC CGAA ACUAGGUG 2685
2993 UGAAACGU c UGCUCCUU 497 AAGGAGCA CUGAUGAG GCCGUUAGGC CGAA ACGUUUCA 2686
2998 CGUCUGCU C CUUGUCCU 498 AGGACAAG CUGAUGAG GCCGUUAGGC CGAA AGCAGACG 2687
3001 CUGCUCCU u GUCCUAAU 499 AUUAGGAC CUGAUGAG GCCGUUAGGC CGAA AGGAGCAG 2688
3004 CUCCUUGU c CUAAUAUU 500 AAUAUUAG CUGAUGAG GCCGUUAGGC CGAA ACAAGGAG 2689 3007 CUUGUCCU A AUAUUCAU 501 AUGAAUAU CUGAUGAG GCCGUUAGGC CGAA AGGACAAG 2690
3010 GUCCUAAU A UUCAUAUC 502 GAUAUGAA CUGAUGAG GCCGUUAGGC CGAA AUUAGGAC 2691
3012 CCUAAUAU U CAUAUCAA 503 UUGAUAUG CUGAUGAG GCCGUUAGGC CGAA AUAUUAGG 2692
3013 CUAAUAUU C AUAUCAAC 504 GUUGAUAU CUGAUGAG GCCGUUAGGC CGAA AAUAUUAG 2693
3016 AUAUUCAU A UCAACAGC 505 GCUGUUGA CUGAUGAG GCCGUUAGGC CGAA AUGAAUAU 2694
3018 AUUCAUAU C AACAGCAC 506 GUGCUGUU CUGAUGAG GCCGUUAGGC CGAA AUAUGAAU 2695
3030 AGCACCAU U CCUGGCAU 507 AUGCCAGG CUGAUGAG GCCGUUAGGC CGAA AUGGUGCU 2696
3031 GCACCAUU C CUGGCAUU 508 AAUGCCAG CUGAUGAG GCCGUUAGGC CGAA AAUGGUGC 2697
3039 CCUGGCAU U CACAUUUU 509 AAAAUGUG CUGAUGAG GCCGUUAGGC CGAA AUGCCAGG 2698
3040 CUGGCAUU C ACAUUUUA 510 UAAAAUGU CUGAUGAG GCCGUUAGGC CGAA AAUGCCAG 2699
3045 AUUCACAU U UUAAAAAU 511 AUUUUUAA CUGAUGAG GCCGUUAGGC CGAA AUGUGAAU 2700
3046 UUCACAUU U UAAAAAUU 512 AAUUUUUA CUGAUGAG GCCGUUAGGC CGAA AAUGUGAA 2701
3047 UCACAUUU u AAAAAUUA 513 UAAUUUUU CUGAUGAG GCCGUUAGGC CGAA AAAUGUGA 2702
3048 CACAUUUU A AAAAUUAU 514 AUAAUUUU CUGAUGAG GCCGUUAGGC CGAA AAAAUGUG 2703
3054 UUAAAAAU U AUGUGGAA 515 UUCCACAU CUGAUGAG GCCGUUAGGC CGAA AUUUUUAA 2704
3055 UAAAAAUU A UGUGGAAG 516 CUUCCACA CUGAUGAG GCCGUUAGGC CGAA AAUUUUUA 2705
3069 AAGUGGAU A GGAGAACU 517 AGUUCUCC CUGAUGAG GCCGUUAGGC CGAA AUCCACUU 2706
3086 GGAGCUGU C AAUAGCCU 518 AGGCUAUU CUGAUGAG GCCGUUAGGC CGAA ACAGCUGC 2707
3090 CUGUCAAU A GCCUAGGG 519 CCCUAGGC CUGAUGAG GCCGUUAGGC CGAA AUUGACAG 2708
3095 AAUAGCCU A GGGCUGAA 520 UUCAGCCC CUGAUGAG GCCGUUAGGC CGAA AGGCUAUU 2709
3105 GGCUGAAU U UUUGUCAG 521 CUGACAAA CUGAUGAG GCCGUUAGGC CGAA AUUCAGCC 2710
3106 GCUGAAUU U UUGUCAGA 522 UCUGACAA CUGAUGAG GCCGUUAGGC CGAA AAUUCAGC 2711
3107 CUGAAUUU U UGUCAGAU 523 AUCUGACA CUGAUGAG GCCGUUAGGC CGAA AAAUUCAG 2712
3108 UGAAUUUU U GUCAGAUA 524 UAUCUGAC CUGAUGAG GCCGUUAGGC CGAA AAAAUUCA 2713
3111 AUUUUUGU C AGAUAAAU 525 AUUUAUCU CUGAUGAG GCCGUUAGGC CGAA ACAAAAAU 2714
3116 UGUCAGAU A AAUAAAAU 526 AUUUUAUU CUGAUGAG GCCGUUAGGC CGAA AUCUGACA 2715
3120 AGAUAAAU A AAAUAAAU 527 AUUUAUUU CUGAUGAG GCCGUUAGGC CGAA AUUUAUCU 2716
3125 AAUAAAAU A AAUCAUUC 528 GAAUGAUU CUGAUGAG GCCGUUAGGC CGAA AUUUUAUU 2717
3129 AAAUAAAU C AUUCAUCC 529 GGAUGAAU CUGAUGAG GCCGUUAGGC CGAA AUUUAUUU 2718
3132 UAAAUCAU U CAUCCUUU 530 AAAGGAUG CUGAUGAG GCCGUUAGGC CGAA AUGAUUUA 2719
3133 AAAUCAUU C AUCCUUUU 531 AAAAGGAU CUGAUGAG GCCGUUAGGC CGAA AAUGAUUU 2720
3136 UCAUUCAU C CUUUUUUU 532 AAAAAAAG CUGAUGAG GCCGUUAGGC CGAA AUGAAUGA 2721
3139 UUCAUCCU U UUUUUGAU 533 AUCAAAAA CUGAUGAG GCCGUUAGGC CGAA AGGAUGAA 2722
3140 UCAUCCUU U UUUUGAUU 534 AAUCAAAA CUGAUGAG GCCGUUAGGC CGAA AAGGAUGA 2723
3141 CAUCCUUU U UUUGAUUA 535 UAAUCAAA CUGAUGAG GCCGUUAGGC CGAA AAAGGAUG 2724
3142 AUCCUUUU U UUGAUUAU 536 AUAAUCAA CUGAUGAG GCCGUUAGGC CGAA AAAAGGAU 2725
3143 UCCUUUUU U UGAUUAUA 537 UAUAAUCA CUGAUGAG GCCGUUAGGC CGAA AAAAAGGA 2726
3144 ccuuuuuu U GAUUAUAA 538 UUAUAAUC CUGAUGAG GCCGUUAGGC CGAA AAAAAAGG 2727
3148 UUUUUGAU U AUAAAAUU 539 AAUUUUAU. .CUGAUGAG. GCCGUUAGGC CGAA AUCAAAAA 2728
3149 UUUUGAUU A UAAAAUUU 540 AAAUUUUA CUGAUGAG GCCGUUAGGC CGAA AAUCAAAA 2729
3151 UUGAUUAU A AAAUUUUC 541 GAAAAUUU CUGAUGAG GCCGUUAGGC CGAA AUAAUCAA 2730
3156 UAUAAAAU U UUCUAAAA 542 UUUUAGAA CUGAUGAG GCCGUUAGGC CGAA AUUUUAUA 2731
3157 AUAAAAUU U UCUAAAAU 543 AUUUUAGA CUGAUGAG GCCGUUAGGC CGAA AAUUUUAU 2732
3158 UAAAAUUU U CUAAAAUG 544 CAUUUUAG CUGAUGAG GCCGUUAGGC CGAA AAAUUUUA 2733
3159 AAAAUUUU C UAAAAUGU 545 ACAUUUUA CUGAUGAG GCCGUUAGGC CGAA AAAAUUUU 2734
3161 AAUUUUCU A AAAUGUAU 546 AUACAUUU CUGAUGAG GCCGUUAGGC CGAA AGAAAAUU 2735
3168 UAAAAUGU A UUUUAGAC 547 GUCUAAAA CUGAUGAG GCCGUUAGGC CGAA ACAUUUUA 2736
3170 AAAUGUAU U UUAGACUU 548 AAGUCUAA CUGAUGAG GCCGUUAGGC CGAA AUACAUUU 2737
3260 AAAUGUAU U UUAGACUU 548 AAGUCUAA CUGAUGAG GCCGUUAGGC CGAA AUACAUUU 2737
3171 AAUGUAUU U UAGACUUC 549 GAAGUCUA CUGAUGAG GCCGUUAGGC CGAA AAUACAUU 2738
3261 AAUGUAUU U UAGACUUC 549 GAAGUCUA CUGAUGAG GCCGUUAGGC CGAA AAUACAUU 2738
3172 AUGUAUUU U AGACUUCC 550 GGAAGUCU CUGAUGAG GCCGUUAGGC CGAA AAAUACAU 2739
3262 AUGUAUUU U AGACUUCC 550 GGAAGUCU CUGAUGAG GCCGUUAGGC CGAA AAAUACAU 2739
3173 UGUAUUUU A GACUUCCU 551 AGGAAGUC CUGAUGAG GCCGUUAGGC CGAA AAAAUACA 2740
3263 UGUAUUUU A GACUUCCU 551 AGGAAGUC CUGAUGAG GCCGUUAGGC CGAA AAAAUACA 2740
3178 UUUAGACU U CCUGUAGG 552 CCUACAGG CUGAUGAG GCCGUUAGGC CGAA AGUCUAAA 2741 3268 UUUAGACU U CCUGUAGG 552 CCUACAGG CUGAUGAG GCCGUUAGGC CGAA AGUCUAAA 2741
3179 UUAGACUU C CUGUAGGG 553 CCCUACAG CUGAUGAG GCCGUUAGGC CGAA AAGUCUAA 2742
3269 UUAGACUU C CUGUAGGG 553 CCCUACAG CUGAUGAG GCCGUUAGGC CGAA AAGUCUAA 2742
3184 CUUCCUGU A GGGGGCGA 554 UCGCCCCC CUGAUGAG GCCGUUAGGC CGAA ACAGGAAG 2743
3274 CUUCCUGU A GGGGGCGA 554 UCGCCCCC CUGAUGAG GCCGUUAGGC CGAA ACAGGAAG 2743
3194 GGGGCGAU A UACUAAAU 555 AUUUAGUA CUGAUGAG GCCGUUAGGC CGAA AUCGCCCC 2744
3247 GGGGCGAU A UACUAAAU 555 AUUUAGUA CUGAUGAG GCCGUUAGGC CGAA AUCGCCCC 2744
3196 GGCGAUAU A CUAAAUGU 556 ACAUUUAG CUGAUGAG GCCGUUAGGC CGAA AUAUCGCC 2745
3249 GGCGAUAU A CUAAAUGU 556 ACAUUUAG CUGAUGAG GCCGUUAGGC CGAA AUAUCGCC 2745
3199 GAUAUACU A AAUGUAUA 557 UAUACAUU CUGAUGAG GCCGUUAGGC CGAA AGUAUAUC 2746
3205 CUAAAUGU A UAUAGUAC 558 GUACUAUA CUGAUGAG GCCGUUAGGC CGAA ACAUUUAG 2747
3207 AAAUGUAU A UAGUACAU 559 AUGUACUA CUGAUGAG GCCGUUAGGC CGAA AUACAUUU 2748
3209 AUGUAUAU A GUACAUUU 560 AAAUGUAC CUGAUGAG GCCGUUAGGC CGAA AUAUACAU 2749
3212 UAUAUAGU A CAUUUAUA 561 UAUAAAUG CUGAUGAG GCCGUUAGGC CGAA ACUAUAUA 2750
3216 UAGUACAU U UAUACUAA 562 UUAGUAUA CUGAUGAG GCCGUUAGGC CGAA AUGUACUA 2751
3217 AGUACAUU U AUAeUAAA 563 UUUAGUAU CUGAUGAG GCCGUUAGGC CGAA AAUGUACU 2752
3218 GUACAUUU A UACUAAAU 564 AUUUAGUA CUGAUGAG GCCGUUAGGC CGAA AAAUGUAC 2753
3220 ACAUUUAU A CUAAAUGU 565 ACAUUUAG CUGAUGAG GCCGUUAGGC CGAA AUAAAUGU 2754
3223 UUUAUACU A AAUGUAUU 566 AAUACAUU CUGAUGAG GCCGUUAGGC CGAA AGUAUAAA 2755
3229 CUAAAUGU A UUCCUGUA 567 UACAGGAA CUGAUGAG GCCGUUAGGC CGAA ACAUUUAG 2756
3231 AAAUGUAU U CCUGUAGG 568 CCUACAGG CUGAUGAG GCCGUUAGGC CGAA AUACAUUU 2757
3232 AAUGUAUU C CUGUAGGG 569 CCCUACAG CUGAUGAG GCCGUUAGGC CGAA AAUACAUU 2758
3237 AUUCCUGU A GGGGGCGA 570 UCGCCCCC CUGAUGAG GCCGUUAGGC CGAA ACAGGAAU 2759
3252 GAUAUACU A AAUGUAUU 571 AAUACAUU CUGAUGAG GCCGUUAGGC CGAA AGUAUAUC 2760
3258 CUAAAUGU A UUUUAGAC 572 GUCUAAAA CUGAUGAG GCCGUUAGGC CGAA ACAUUUAG 2761
3284 GGGGCGAU A AAAUAAAA 573 UUUUAUUU CUGAUGAG GCCGUUAGGC CGAA AUCGCCCC 2762
3289 GAUAAAAU A AAAUGCUA 574 UAGCAUUU CUGAUGAG GCCGUUAGGC CGAA AUUUUAUC 2763
3297 AAAAUGCU A AACAACUG 575 CAGUUGUU CUGAUGAG GCCGUUAGGC CGAA AGCAUUUU 2764
Input Sequence = NM_001285. Cut Site = UH/.
Arm Length = 8. Core Sequence = CUGAUGAG GCCGUUAGGC CGAA Underlined region can be any X sequence or linker, as described herein. NM_001285 (Homo sapiens chloride channel, calcium activated, 1 (CLCAl) mRNA, 3311 bp)
Table IV: Human CLCAl Inozyme and Target Sequence 249.021
Pos Substrate Seq ID Inozyme Rz
No. Seq ID
No.
10 GCUAAUGC U UUUGGUAC 576 GUACCAAA CUGAUGAG GCCGUUAGGC CGAA ICAUUAGC 2765
19 UUUGGUAC A AAUGGAUG 577 CAUCCAUU CUGAUGAG GCCGUUAGGC CGAA IUACCAAA 2766
50 AUAUUUUC U UGUUUAAG 578 CUUAAACA CUGAUGAG GCCGUUAGGC CGAA IAAAAUAU 2767
65 AGGGGAGC A UGAAGAGG 579 CCUCUUCA CUGAUGAG GCCGUUAGGC CGAA ICUCCCCU 2768
89 GUUAUGUC A AGCAUCUG 580 CAGAUGCU CUGAUGAG GCCGUUAGGC CGAA IACAUAAC 2769
93 UGUCAAGC A UCUGGCAC 581 GUGCCAGA CUGAUGAG GCCGUUAGGC CGAA ICUUGACA 2770
96 CAAGCAUC U GGCACAGC 582 GCUGUGCC CUGAUGAG GCCGUUAGGC CGAA IAUGCUUG 2771
100 CAUCUGGC A CAGCUGAA 583 UUCAGCUG CUGAUGAG GCCGUUAGGC CGAA ICCAGAUG 2772
102 UCUGGCAC A GCUGAAGG 584 CCUUCAGC CUGAUGAG GCCGUUAGGC CGAA IUGCCAGA 2773
105 GGCACAGC U GAAGGCAG 585 CUGCCUUC CUGAUGAG GCCGUUAGGC CGAA ICUGUGCC 2774
112 CUGAAGGC A GAUGGAAA 586 UUUCCAUC CUGAUGAG GCCGUUAGGC CGAA ICCUUCAG 2775
128 AUAUUUAC A AGUACGCA 587 UGCGUACU CUGAUGAG GCCGUUAGGC CGAA IUAAAUAU 2776
136 AAGUACGC A AUUUGAGA 588 UCUCAAAU CUGAUGAG GCCGUUAGGC CGAA ICGUACUU 2777
146 UUUGAGAC U AAGAUAUU 589 AAUAUCUϋ CUGAUGAG GCCGUUAGGC CGAA IUCUCAAA 2778
161 UUGUUAUC A UUCUCCUA 590 UAGGAGAA CUGAUGAG GCCGUUAGGC CGAA IAUAACAA 2779
165 UAUCAUUC U CCUAUUGA 591 UCAAUAGG CUGAUGAG GCCGUUAGGC CGAA IAAUGAUA 2780
167 UCAUUCUC C UAUUGAAG 592 CUUCAAUA CUGAUGAG GCCGUUAGGC CGAA IAGAAUGA 2781
168 CAUUCUCC U AUUGAAGA 593 UCUUCAAU CUGAUGAG GCCGUUAGGC CGAA IGAGAAUG 2782
178 UUGAAGAC A AGAGCAAU 594 AUUGCUCU CUGAUGAG GCCGUUAGGC CGAA IUCUUCAA 2783
184 ACAAGAGC A AUAGUAAA 595 UUUACUAU CUGAUGAG GCCGUUAGGC CGAA ICUCUUGU 2784
195 AGUAAAAC A CAUCAGGU 596 ACCUGAUG CUGAUGAG GCCGUUAGGC CGAA IUUUUACU 2785
197 UAAAACAC A UCAGGUCA 597 UGACCUGA CUGAUGAG GCCGUUAGGC CGAA IUGUUUUA 2786
200 AACACAUC A GGUCAGGG 598 CCCUGACC CUGAUGAG GCCGUUAGGC CGAA IAUGUGUU 2787
205 AUCAGGUC A GGGGGUUA 599 UAACCCCC CUGAUGAG GCCGUUAGGC CGAA IACCUGAU 2788
219 UUAAAGAC C UGUGAUAA 600 UUAUCACA CUGAUGAG GCCGUUAGGC CGAA IUCUUUAA 2789
220 UAAAGACC u GUGAUAAA 601 UUUAUCAC CUGAUGAG GCCGUUAGGC CGAA IGUCUUUA 2790
230 UGAUAAAC c ACUUCCGA 602 UCGGAAGU CUGAUGAG GCCGUUAGGC CGAA IUUUAUCA 2791
231 GAUAAACC A CUUCCGAU 603 AUCGGAAG CUGAUGAG GCCGUUAGGC CGAA IGUUUAUC 2792
233 UAAACCAC U UCCGAUAA 604 UUAUCGGA CUGAUGAG GCCGUUAGGC CGAA lUGGUUUA 2793
236 ACCACUUC C GAUAAGUU 605 AACUUAUC CUGAUGAG GCCGUUAGGC CGAA lAAGUGGU 2794
258 CGUGUGUC U AUAUUUUC 606 GAAAAUAU CUGAUGAG GCCGUUAGGC CGAA IACACACG 2795
267 AUAUUUUC A UAUCUGUA 607 UACAGAUA CUGAUGAG GCCGUUAGGC CGAA lAAAAUAU 2796
272 UUCAUAUC U GUAUAUAU 608 AUAUAUAC CUGAUGAG GCCGUUAGGC CGAA IAUAUGAA 2797
299 AGAAAGAC A CCUUCGUA 609 UACGAAGG CUGAUGAG GCCGUUAGGC CGAA IUCUUUCU 2798
301 AAAGACAC C UUCGUAAC 610 GUUACGAA CUGAUGAG GCCGUUAGGC CGAA IUGUCUUU 2799
302 AAGACACC U UCGUAACC 611 GGUUACGA CUGAUGAG GCCGUUAGGC CGAA IGUGUCUU 2800
310 UUCGUAAC C CGCAUUUU 612 AAAAUGCG CUGAUGAG GCCGUUAGGC CGAA IUUACGAA 2801
311 UCGUAACC C GCAUUUUC 613 GAAAAUGC CUGAUGAG GCCGUUAGGC CGAA IGUUACGA 2802
314 UAACCCGC A UUUUCCAA 614 UUGGAAAA CUGAUGAG GCCGUUAGGC CGAA ICGGGUUA 2803
320 GCAUUUUC C AAAGAGAG 615 CUCUCUUU CUGAUGAG GCCGUUAGGC CGAA lAAAAUGC 2804
321 CAUUUUCC A AAGAGAGG 616 CCUCUCUU CUGAUGAG GCCGUUAGGC CGAA IGAAAAUG 2805
334 GAGGAAUC A CAGGGAGA 617 UCUCCCUG CUGAUGAG GCCGUUAGGC CGAA IAUUCCUC 2806
336 GGAAUCAC A GGGAGAUG 618 CAUCUCCC CUGAUGAG GCCGUUAGGC CGAA IUGAUUCC 2807
348 AGAUGUAC A GCAAUGGG 619 CCCAUUGC CUGAUGAG GCCGUUAGGC CGAA IUACAUCU 2808
351 UGUACAGC A AUGGGGCC 620 GGCCCCAU CUGAUGAG GCCGUUAGGC CGAA ICUGUACA 2809
359 AAUGGGGC C AUUUAAGA 621 UCUUAAAU CUGAUGAG GCCGUUAGGC CGAA ICCCCAUU 2810
360 AUGGGGCC A UUUAAGAG 622 CUCUUAAA CUGAUGAG GCCGUUAGGC CGAA IGCCCCAU 2811
372 AAGAGUUC U GUGUUCAU 623 AUGAACAC CUGAUGAG GCCGUUAGGC CGAA lAACUCUU 2812
379 CUGUGUUC A UCUUGAUU 624 AAUCAAGA CUGAUGAG GCCGUUAGGC CGAA IAACACAG 2813
382 UGUUCAUC U UGAUUCUU 625 AAGAAUCA CUGAUGAG GCCGUUAGGC CGAA lAUGAACA 2814
389 CUUGAUUC U UCACCUUC 626 GAAGGUGA CUGAUGAG GCCGUUAGGC CGAA IAAUCAAG 2815
392 GAUUCUUC A CCUUCUAG 627 CUAGAAGG CUGAUGAG GCCGUUAGGC CGAA IAAGAAUC 2816
394 UUCUUCAC C UUCUAGAA 628 UUCUAGAA CUGAUGAG GCCGUUAGGC CGAA lUGAAGAA 2817
395 UCUUCACC U UCUAGAAG 629 CUUCUAGA CUGAUGAG GCCGUUAGGC CGAA IGUGAAGA 2818
398 UCACCUUC U AGAAGGGG 630 CCCCUUCU CUGAUGAG GCCGUUAGGC CGAA IAAGGUGA 2819
408 GAAGGGGC c CUGAGUAA 631 UUACUCAG CUGAUGAG GCCGUUAGGC CGAA ICCCCUUC 2820
409 AAGGGGCC c UGAGUAAU 632 AUUACUCA CUGAUGAG GCCGUUAGGC CGAA IGCCCCUU 2821
410 AGGGGCCC u GAGUAAUU 633 AAUUACUC CUGAUGAG GCCGUUAGGC CGAA IGGCCCCU 2822
420 AGUAAUUC A CUCAUUCA 634 UGAAUGAG CUGAUGAG GCCGUUAGGC CGAA IAAUUACU 2823
422 UAAUUCAC U CAUUCAGC 635 GCUGAAUG CUGAUGAG GCCGUUAGGC CGAA IUGAAUUA 2824
424 AUUCACUC A UUCAGCUG 636 CAGCUGAA CUGAUGAG GCCGUUAGGC CGAA IAGUGAAU 2825
428 ACUCAUUC A GCUGAACA 637 UGUUCAGC CUGAUGAG GCCGUUAGGC CGAA IAAUGAGU 2826
431 CAUUCAGC U GAACAACA 638 UGUUGUUC CUGAUGAG GCCGUUAGGC CGAA ICUGAAUG 2827
436 AGCUGAAC A ACAAUGGC 639 GCCAUUGU CUGAUGAG GCCGUUAGGC CGAA IUUCAGCU 2828
439 UGAACAAC A AUGGCUAU 640 AUAGCCAU CUGAUGAG GCCGUUAGGC CGAA IUUGUUCA 2829
445 ACAAUGGC U AUGAAGGC 641 GCCUUCAU CUGAUGAG GCCGUUAGGC CGAA ICCAUUGU 2830
454 AUGAAGGC A UUGUCGUU 642 AACGACAA CUGAUGAG GCCGUUAGGC CGAA ICCUUCAU 2831
465 GUCGUUGC A AUCGACCC 643 GGGUCGAU CUGAUGAG GCCGUUAGGC CGAA ICAACGAC 2832
472 CAAUCGAC C CCAAUGUG 644 CACAUUGG CUGAUGAG GCCGUUAGGC CGAA IUCGAUUG 2833
473 AAUCGACC c CAAUGUGC 645 GCACAUUG CUGAUGAG GCCGUUAGGC CGAA IGUCGAUU 2834
474 AUCGACCC c AAUGUGCC 646 GGCACAUϋ CUGAUGAG GCCGUUAGGC CGAA IGGUCGAU 2835
475 UCGACCCC A AUGUGCCA 647 UGGCACAU CUGAUGAG GCCGUUAGGC CGAA IGGGUCGA 2836
482 CAAUGUGC C AGAAGAUG 648 CAUCUUCU CUGAUGAG GCCGUUAGGC CGAA ICACAUUG 2837
483 AAUGUGCC A GAAGAUGA 649 UCAUCUUC CUGAUGAG GCCGUUAGGC CGAA IGCACAUU 2838
495 GAUGAAAC A CUCAUUCA 650 UGAAUGAG CUGAUGAG GCCGUUAGGC CGAA IUUUCAUC 2839
497 UGAAACAC U CAUUCAAC 651 GUUGAAUG CUGAUGAG GCCGUUAGGC CGAA IUGUUUCA 2840
499 AAACACUC A UUCAACAA 652 UUGUUGAA CUGAUGAG GCCGUUAGGC CGAA IAGUGUUU 2841
503 ACUCAUUC A ACAAAUAA 653 UUAUUUGU CUGAUGAG GCCGUUAGGC CGAA IAAUGAGU 2842
506 CAUUCAAC A AAUAAAGG 654 CCUUUAUU CUGAUGAG GCCGUUAGGC CGAA IUUGAAUG 2843
517 UAAAGGAC A UGGUGACC 655 GGUCACCA CUGAUGAG GCCGUUAGGC CGAA IUCCUUUA 2844
525 AUGGUGAC C CAGGCAUC 656 GAUGCCUG CUGAUGAG GCCGUUAGGC CGAA IUCACCAU 2845
526 UGGUGACC C AGGCAUCU 657 AGAUGCCU CUGAUGAG GCCGUUAGGC CGAA IGUCACCA 2846
527 GGUGACCC A GGCAUCUC 658 GAGAUGCC CUGAUGAG GCCGUUAGGC CGAA IGGUCACC 2847
531 ACCCAGGC A UCUCUGUA 659 UACAGAGA CUGAUGAG GCCGUUAGGC CGAA ICCUGGGU 2848
534 CAGGCAUC U CUGUAUCU 660 AGAUACAG CUGAUGAG GCCGUUAGGC CGAA IAUGCCUG 2849
536 GGCAUCUC U GUAUCUGU 661 • ACAGAUAC •CUGAUGAG GCCGUUAGGC CGAA IAGAUGCC 2850
542 UCUGUAUC U GUUUGAAG 662 CUUCAAAC CUGAUGAG GCCGUUAGGC CGAA IAUACAGA 2851
552 UUUGAAGC u ACAGGAAA 663 UUUCCUGU CUGAUGAG GCCGUUAGGC CGAA ICUUCAAA 2852
555 GAAGCUAC A GGAAAGCG 664 CGCUUUCC CUGAUGAG GCCGUUAGGC CGAA IUAGCUUC 2853
574 UUUAUUUC A AAAAUGUU 665 AACAUUUU CUGAUGAG GCCGUUAGGC CGAA IAAAUAAA 2854
585 AAUGUUGC C AUUUUGAU 666 AUCAAAAU CUGAUGAG GCCGUUAGGC CGAA ICAACAUU 2855
586 AUGUUGCC A UUUUGAUU 667 AAUCAAAA CUGAUGAG GCCGUUAGGC CGAA IGCAACAU 2856
596 UUUGAUUC C UGAAACAU 668 AUGUUUCA CUGAUGAG GCCGUUAGGC CGAA IAAUCAAA 2857
597 UUGAUUCC U GAAACAUG 669 CAUGUUUC CUGAUGAG GCCGUUAGGC CGAA IGAAUCAA 2858
603 CCUGAAAC A UGGAAGAC 670 GUCUUCCA CUGAUGAG GCCGUUAGGC CGAA IUUUCAGG 2859
612 UGGAAGAC A AAGGCUGA 671 UCAGCCUU CUGAUGAG GCCGUUAGGC CGAA lUCUUCCA 2860
618 ACAAAGGC U GACUAUGU 672 ACAUAGUC CUGAUGAG GCCGUUAGGC CGAA ICCUUUGU 2861
622 AGGCUGAC U AUGUGAGA 673 UCUCACAU CUGAUGAG GCCGUUAGGC CGAA IUCAGCCU 2862
632 UGUGAGAC C AAAACUUG 674 CAAGUUUU CUGAUGAG GCCGUUAGGC CGAA IUCUCACA 2863
633 GUGAGACC A AAACUUGA 675 UCAAGUUU CUGAUGAG GCCGUUAGGC CGAA IGUCUCAC 2864
638 ACCAAAAC U UGAGACCU 676 AGGUCUCA CUGAUGAG GCCGUUAGGC CGAA IUUUUGGU 2865
645 CUUGAGAC C UACAAAAA 677 UUUUUGUA CUGAUGAG GCCGUUAGGC CGAA IUCUCAAG 2866
646 UUGAGACC U ACAAAAAU 678 AUUUUUGU CUGAUGAG GCCGUUAGGC CGAA IGUCUCAA 2867
649 AGACCUAC A AAAAUGCU 679 AGCAUUUU CUGAUGAG GCCGUUAGGC CGAA lUAGGUCU 2868
657 AAAAAUGC U GAUGUUCU 680 AGAACAUC CUGAUGAG GCCGUUAGGC CGAA ICAUUUUU 2869
665 UGAUGUUC U GGUUGCUG 681 CAGCAACC CUGAUGAG GCCGUUAGGC CGAA IAACAUCA 2870
672 CUGGUUGC u GAGUCUAC 682 GUAGACUC •CUGAUGAG GCCGUUAGGC CGAA ICAACCAG 2871
678 GCUGAGUC u ACUCCUCC 683 GGAGGAGU CUGAUGAG GCCGUUAGGC CGAA IACUCAGC 2872
681 GAGUCUAC u CCUCCAGG 684 CCUGGAGG CUGAUGAG GCCGUUAGGC CGAA IUAGACUC 2873
683 GUCUACUC c UCCAGGUA 685 UACCUGGA CUGAUGAG GCCGUUAGGC CGAA IAGUAGAC 2874
684 UCUACUCC u CCAGGUAA 686 UUACCUGG CUGAUGAG GCCGUUAGGC CGAA IGAGUAGA 2875
686 UACUCCUC c AGGUAAUG 687 CAUUACCU CUGAUGAG GCCGUUAGGC CGAA IAGGAGUA 2876
687 ACUCCUCC A GGUAAUGA 688 UCAUUACC CUGAUGAG GCCGUUAGGC CGAA IGAGGAGU 2877
701 UGAUGAAC c CUACACUG 689 CAGUGUAG CUGAUGAG GCCGUUAGGC CGAA IUUCAUCA 2878
702 GAUGAACC c UACACUGA 690 UCAGUGUA CUGAUGAG GCCGUUAGGC CGAA IGUUCAUC 2879
703 AUGAACCC u ACACUGAG 691 CUCAGUGU CUGAUGAG GCCGUUAGGC CGAA IGGUUCAU 2880
706 AACCCUAC A CUGAGCAG 692 CUGCUCAG CUGAUGAG GCCGUUAGGC CGAA IUAGGGUU 2881
708 CCCUACAC U GAGCAGAU 693 AUCUGCUC CUGAUGAG GCCGUUAGGC CGAA IUGUAGGG 2882
713 CACUGAGC A GAUGGGCA 694 UGCCCAUC CUGAUGAG GCCGUUAGGC CGAA ICUCAGUG 2883
721 AGAUGGGC A ACUGUGGA 695 UCCACAGU CUGAUGAG GCCGUUAGGC CGAA ICCCAUCU 2884
724 UGGGCAAC U GUGGAGAG 696 CUCUCCAC CUGAUGAG GCCGUUAGGC CGAA IUUGCCCA 2885
748 AAAGGAUC C ACCUCACU 697 AGUGAGGU CUGAUGAG GCCGUUAGGC CGAA IAUCCUUU 2886
749 AAGGAUCC A CCUCACUC 698 GAGUGAGG CUGAUGAG GCCGUUAGGC CGAA IGAUCCUU 2887
751 GGAUCCAC C UCACUCCU 699 AGGAGUGA CUGAUGAG GCCGUUAGGC CGAA IUGGAUCC 2888
752 GAUCCACC u CACUCCUG 700 CAGGAGUG CUGAUGAG GCCGUUAGGC CGAA IGUGGAUC 2889
754 UCCACCUC A CUCCUGAU 701 AUCAGGAG CUGAUGAG GCCGUUAGGC CGAA lAGGUGGA 2890
756 CACCUCAC U CCUGAUUU 702 AAAUCAGG CUGAUGAG GCCGUUAGGC CGAA IUGAGGUG 2891
758 CCUCACUC C UGAUUUCA 703 UGAAAUCA CUGAUGAG GCCGUUAGGC CGAA IAGUGAGG 2892
759 CUCACUCC u GAUUUCAU 704 AUGAAAUC CUGAUGAG GCCGUUAGGC CGAA IGAGUGAG 2893
766 CUGAUUUC A UUGCAGGA 705 UCCUGCAA CUGAUGAG GCCGUUAGGC CGAA IAAAUCAG 2894
771 UUCAUUGC A GGAAAAAA 706 UUUUUUCC CUGAUGAG GCCGUUAGGC CGAA ICAAUGAA 2895
786 AAGUUAGC U GAAUAUGG 707 CCAUAUUC CUGAUGAG GCCGUUAGGC CGAA ICUAACUU 2896
797 AUAUGGAC C ACAAGGUA 708 UACCUUGU CUGAUGAG GCCGUUAGGC CGAA IUCCAUAU 2897
798 UAUGGACC A CAAGGUAA 709 UUACCUUG CUGAUGAG GCCGUUAGGC CGAA IGUCCAUA 2898
800 UGGACCAC A AGGUAAGG 710 CCUUACCU CUGAUGAG GCCGUUAGGC CGAA IUGGUCCA 2899
810 GGUAAGGC A UUUGUCCA 711 UGGACAAA CUGAUGAG GCCGUUAGGC CGAA ICCUUACC 2900
817 CAUUUGUC C AUGAGUGG 712 CCACUCAU CUGAUGAG GCCGUUAGGC CGAA IACAAAUG 2901
818 AUUUGUCC A UGAGUGGG 713 CCCACUCA CUGAUGAG GCCGUUAGGC CGAA IGACAAAU 2902
828 GAGUGGGC U CAUCUACG 714 CGUAGAUG CUGAUGAG GCCGUUAGGC CGAA ICCCACUC 2903
830 GUGGGCUC A UCUACGAU 715 AUCGUAGA CUGAUGAG GCCGUUAGGC CGAA IAGCCCAC 2904
833 GGCUCAUC U ACGAUGGG 716 CCCAUCGU CUGAUGAG GCCGUUAGGC CGAA IAUGAGCC 2905
859 ACGAGUAC A AUAAUGAU 717 AUCAUUAU CUGAUGAG GCCGUUAGGC CGAA IUACUCGU 2906
877 AGAAAUUC U ACUUAUCC 718 GGAUAAGU CUGAUGAG GCCGUUAGGC CGAA IAAUUUCU 2907
880 AAUUCUAC U UAUCCAAU 719 AUUGGAUA CUGAUGAG GCCGUUAGGC CGAA IUAGAAUU 2908
885 UACUUAUC C AAUGGAAG 720 CUUCCAUU CUGAUGAG GCCGUUAGGC CGAA IAUAAGUA 2909
886 ACUUAUCC A AUGGAAGA 721 UCUUCCAU CUGAUGAG GCCGUUAGGC CGAA IGAUAAGU 2910
899 AAGAAUAC A AGCAGUAA 722 UUACUGCU CUGAUGAG GCCGUUAGGC CGAA IUAUUCUU 2911
903 AUACAAGC A GUAAGAUG 723 CAUCUUAC CUGAUGAG GCCGUUAGGC CGAA ICUUGUAU 2912
915 AGAUGUUC A GCAGGUAU 724 AUACCUGC CUGAUGAG GCCGUUAGGC CGAA lAACAUCU 2913
918 UGUUCAGC A GGUAUUAC 725 GUAAUACC CUGAUGAG GCCGUUAGGC CGAA ICUGAACA 2914
927 GGUAUUAC U GGUACAAA 726 UUUGUACC CUGAUGAG GCCGUUAGGC CGAA lUAAUACC 2915
933 ACUGGUAC A AAUGUAGU 727 ACUACAUU CUGAUGAG GCCGUUAGGC CGAA IUACCAGU 2916
953 GAAGUGUC A GGGAGGCA 728 UGCCUCCC CUGAUGAG GCCGUUAGGC CGAA IACACUUC 2917
961 AGGGAGGC A GCUGUUAC 729 GUAACAGC CUGAUGAG GCCGUUAGGC CGAA ICCUCCCU 2918
964 GAGGCAGC U GUUACACC 730 GGUGUAAC CUGAUGAG GCCGUUAGGC CGAA ICUGCCUC 2919
970 GCUGUUAC A CCAAAAGA 731 UCUUUUGG CUGAUGAG GCCGUUAGGC CGAA IUAACAGC 2920
972 UGUUACAC C AAAAGAUG 732 CAUCUUUU CUGAUGAG GCCGUUAGGC CGAA IUGUAACA 2921
973 GUUACACC A AAAGAUGC 733 GCAUCUUU CUGAUGAG GCCGUUAGGC CGAA IGUGUAAC 2922
982 AAAGAUGC A CAUUCAAU 734 AUUGAAUG CUGAUGAG GCCGUUAGGC CGAA ICAUCUUU 2923
984 AGAUGCAC A UUCAAUAA 735 UUAUUGAA CUGAUGAG GCCGUUAGGC CGAA lUGCAUCU 2924
988 GCACAUUC A AUAAAGUU 736 AACUUUAU CUGAUGAG GCCGUUAGGC CGAA IAAUGUGC 2925
999 AAAGUUAC A GGACUCUA 737 UAGAGUCC CUGAUGAG GCCGUUAGGC CGAA lUAACUUU 2926
1004 UACAGGAC U CUAUGAAA 738 UUUCAUAG CUGAUGAG GCCGUUAGGC CGAA lUCCUGUA 2927
1006 CAGGACUC U AUGAAAAA 739 UUUUUCAU CUGAUGAG GCCGUUAGGC CGAA IAGUCCUG 2928
1031 GUUUGUUC U CCAAUCCC 740 GGGAUUGG CUGAUGAG GCCGUUAGGC CGAA IAACAAAC 2929
1033 UUGUUCUG C AAUCCCGC 741 GCGGGAUU CUGAUGAG GCCGUUAGGC CGAA IAGAACAA 2930
1034 UGUUCUCC A AUCCCGCC 742 GGCGGGAU CUGAUGAG GCCGUUAGGC CGAA IGAGAACA 2931
1038 CUCCAAUC C CGCCAGAC 743 GUCUGGCG CUGAUGAG GCCGUUAGGC CGAA IAUUGGAG 2932
1039 UCCAAUCC C GCCAGACG 744 CGUCUGGC CUGAUGAG GCCGUUAGGC CGAA IGAUUGGA 2933
1042 AAUCCCGC c AGACGGAG 745 CUCCGUCU CUGAUGAG GCCGUUAGGC CGAA ICGGGAUU 2934
1043 AUCCCGCC A GACGGAGA 746 UCUCCGUC CUGAUGAG GCCGUUAGGC CGAA IGCGGGAU 2935
1056 GAGAAGGC U UCUAUAAU 747 AUUAUAGA CUGAUGAG GCCGUUAGGC CGAA ICCUUCUC 2936
1059 AAGGCUUC U AUAAUGUU 748 AACAUUAU CUGAUGAG GCCGUUAGGC CGAA IAAGCCUU 2937
1071 AUGUUUGC A CAACAUGU 749 ACAUGUUG CUGAUGAG GCCGUUAGGC CGAA ICAAACAU 2938
1073 GUUUGCAC A ACAUGUUG 750 CAACAUGU CUGAUGAG GCCGUUAGGC CGAA IUGCAAAC 2939
1076 UGCACAAC A UGUUGAUU 751 AAUCAACA CUGAUGAG GCCGUUAGGC CGAA IUUGUGCA 2940
1086 GUUGAUUC U AUAGUUGA 752 UCAACUAU CUGAUGAG GCCGUUAGGC CGAA IAAUCAAC 2941
1099 UUGAAUUC U GUACAGAA 753 UUCUGUAC CUGAUGAG GCCGUUAGGC CGAA IAAUUCAA 2942
1104 UUCUGUAC A GAACAAAA 754 UUUUGUUC CUGAUGAG GCCGUUAGGC CGAA IUACAGAA 2943
1109 UACAGAAC A AAACCACA 755 UGUGGUUU CUGAUGAG GCCGUUAGGC CGAA IUUCUGUA 2944
1114 AACAAAAC C ACAACAAA 756 UUUGUUGU CUGAUGAG GCCGUUAGGC CGAA IUUUUGUU 2945
1115 ACAAAACC A CAACAAAG 757 CUUUGUUG CUGAUGAG GCCGUUAGGC CGAA IGUUUUGU 2946
1117 AAAACCAC A ACAAAGAA 758 UUCUUUGU CUGAUGAG GCCGUUAGGC CGAA IUGGUUUU 2947
1120 ACCACAAC A AAGAAGCU 759 AGCUUCUU CUGAUGAG GCCGUUAGGC CGAA IUUGUGGU 2948
1128 AAAGAAGC U CCAAACAA 760 UUGUUUGG CUGAUGAG GCCGUUAGGC CGAA ICUUCUUU 2949
1130 AGAAGCUC C AAACAAGC 761 GCUUGUUU CUGAUGAG GCCGUUAGGC CGAA IAGCUUCU 2950
1131 GAAGCUCC A AACAAGCA 762 UGCUUGUU CUGAUGAG GCCGUUAGGC CGAA IGAGCUUC 2951
1135 CUCCAAAC A AGCAAAAU 763 AUUUUGCU CUGAUGAG GCCGUUAGGC CGAA IUUUGGAG 2952
1139 AAACAAGC A AAAUCAAA 764 UUUGAUUU CUGAUGAG GCCGUUAGGC CGAA ICUUGUUU 2953
1145 GCAAAAUC A AAAAUGCA 765 UGCAUUUU CUGAUGAG GCCGUUAGGC CGAA IAUUUUGC 2954
1153 AAAAAUGC A AUCUCCGA 766 UCGGAGAU CUGAUGAG GCCGUUAGGC CGAA ICAUUUUU 2955
1157 AUGCAAUC U CCGAAGCA 767 UGCUUCGG CUGAUGAG GCCGUUAGGC CGAA lAUUGCAU 2956
1159 GCAAUCUC C GAAGCACA 768 UGUGCUUC CUGAUGAG GCCGUUAGGC CGAA IAGAUUGC 2957
1165 UCCGAAGC A CAUGGGAA 769 UUCCCAUG CUGAUGAG GCCGUUAGGC CGAA ICUUCGGA 2958
1167 CGAAGCAC A UGGGAAGU 770 ACUUCCCA CUGAUGAG GCCGUUAGGC CGAA IUGCUUCG 2959
1180 AAGUGAUC C GUGAUUCU 771 AGAAUCAC CUGAUGAG GCCGUUAGGC CGAA IAUCACUU 2960
1188 CGUGAUUC U GAGGACUU 772 AAGUCCUC CUGAUGAG GCCGUUAGGC CGAA IAAUCACG 2961
1195 CUGAGGAC u UUAAGAAA 773 UUUCUUAA CUGAUGAG GCCGUUAGGC CGAA IUCCUCAG 2962
1206 AAGAAAAC c ACUCCUAU 774 AUAGGAGU CUGAUGAG GCCGUUAGGC CGAA lUUUUCUU 2963
1207 AGAAAACC A CUCCUAUG 775 CAUAGGAG CUGAUGAG GCCGUUAGGC CGAA IGUUUUCU 2964
1209 AAAACCAC U CCUAUGAC 776 GUCAUAGG CUGAUGAG GCCGUUAGGC CGAA IUGGUUUU 2965
1211 AACCACUC C UAUGACAA 777 UUGUCAUA CUGAUGAG GCCGUUAGGC CGAA IAGUGGUU 2966
1212 ACCACUCC U AUGACAAC 778 GUUGUCAU CUGAUGAG GCCGUUAGGC CGAA IGAGUGGU 2967
1218 CCUAUGAC A ACACAGCC 779 GGCUGUGU CUGAUGAG GCCGUUAGGC CGAA IUCAUAGG 2968
1221 AUGACAAC A CAGCCACC 780 GGUGGCUG CUGAUGAG GCCGUUAGGC CGAA IUUGUCAU 2969
1223 GACAACAC A GCCACCAA 781 UUGGUGGC CUGAUGAG GCCGUUAGGC CGAA IUGUUGUC 2970
1226 AACACAGC C ACCAAAUC 782 GAUUUGGU CUGAUGAG GCCGUUAGGC CGAA ICUGUGUU 2971
1227 ACACAGCC A CCAAAUCC 783 GGAUUUGG CUGAUGAG GCCGUUAGGC CGAA IGCUGUGU 2972
1229 ACAGCCAC C AAAUCCCA 784 UGGGAUUU CUGAUGAG GCCGUUAGGC CGAA IUGGCUGU 2973
1230 CAGCCACC A AAUCCCAC 785 GUGGGAUU CUGAUGAG GCCGUUAGGC CGAA IGUGGCUG 2974
1235 ACCAAAUC C CACCUUCU 786 AGAAGGUG CUGAUGAG GCCGUUAGGC CGAA.IAUUUGGU 2975
1236 CCAAAUCC C ACCUUCUC 787^ GAGAAGGU CUGAUGAG GCCGUUAGGC CGAA IGAUUUGG 2976
1237 CAAAUCCC A CCUUCUCA 788 UGAGAAGG CUGAUGAG GCCGUUAGGC CGAA IGGAUUUG 2977
1239 AAUCCCAC C UUCUCAUU 789 AAUGAGAA CUGAUGAG GCCGUUAGGC CGAA IUGGGAUU 2978
1240 AUCCCACC u UCUCAUUG 790 CAAUGAGA CUGAUGAG GCCGUUAGGC CGAA IGUGGGAU 2979
1243 CCACCUUC u CAUUGCUG 791 CAGCAAUG CUGAUGAG GCCGUUAGGC CGAA -IAAGGUGG 2980
1245 ACCUUCUC A UUGCUGCA 792 UGCAGCAA CUGAUGAG GCCGUUAGGC CGAA IAGAAGGU 2981
1250 CUCAUUGC U GCAGAUUG 793 CAAUCUGC CUGAUGAG GCCGUUAGGC CGAA ICAAUGAG 2982
1253 AUUGCUGC A GAUUGGAC 794 GUCCAAUC CUGAUGAG GCCGUUAGGC CGAA ICAGCAAU 2983
1262 GAUUGGAC A AAGAAUUG 795 CAAUUCUU CUGAUGAG GCCGUUAGGC CGAA IUCCAAUC 2984
1282 GUUUAGUC C UUGACAAA 796 UUUGUCAA CUGAUGAG GCCGUUAGGC CGAA IACUAAAC 2985
1283 UUUAGUCC U UGACAAAU 797 AUUUGUCA CUGAUGAG GCCGUUAGGC CGAA IGACUAAA 2986
1288 UCCUUGAC A AAUCUGGA 798 UCCAGAUU CUGAUGAG GCCGUUAGGC CGAA IUCAAGGA 2987
1293 GACAAAUC U GGAAGCAU 799 AUGCUUCC CUGAUGAG GCCGUUAGGC CGAA IAUUUGUC 2988
1300 CUGGAAGC A UGGCGACU 800 AGUCGCCA CUGAUGAG GCCGUUAGGC CGAA ICUUCCAG 2989
1308 AUGGCGAC U GGUAACCG 801 CGGUUACC CUGAUGAG GCCGUUAGGC CGAA IUCGCCAU 2990
1315 CUGGUAAC C GCCUCAAU 802 AUUGAGGC CUGAUGAG GCCGUUAGGC CGAA lUUACCAG 2991
1318 GUAACCGC C UCAAUCGA 803 UCGAUUGA CUGAUGAG GCCGUUAGGC CGAA ICGGUUAC 2992
1319 UAACCGCC U CAAUCGAC 804 GUCGAUUG CUGAUGAG GCCGUUAGGC CGAA IGCGGUUA 2993
1321 ACCGCCUC A AUCGACUG 805 CAGUCGAU CUGAUGAG GCCGUUAGGC CGAA IAGGCGGU 2994
1328 CAAUCGAC U GAAUCAAG 806 CUUGAUUC CUGAUGAG GCCGUUAGGC CGAA IUCGAUUG 2995
1334 ACUGAAUC A AGCAGGCC 807 GGCCUGCU CUGAUGAG GCCGUUAGGC CGAA IAUUCAGU 2996
1338 AAUCAAGC A GGCCAGCU 808 AGCUGGCC CUGAUGAG GCCGUUAGGC CGAA ICUUGAUU 2997
1342 AAGCAGGC C AGCUUUUC 809 GAAAAGCU CUGAUGAG GCCGUUAGGC CGAA ICCUGCUU 2998
1343 AGCAGGCC A GCUUUUCC 810 GGAAAAGC CUGAUGAG GCCGUUAGGC CGAA IGCCUGCU 2999
1346 AGGCCAGC U UUUCCUGC 811 GCAGGAAA CUGAUGAG GCCGUUAGGC CGAA ICUGGCCU 3000
1351 AGCUUUUC C UGCUGCAG 812 CUGCAGCA CUGAUGAG GCCGUUAGGC CGAA IAAAAGCU 3001
1352 GCUUUUCC U GCUGCAGA 813 UCUGCAGC CUGAUGAG GCCGUUAGGC CGAA IGAAAAGC 3002
1355 UUUCCUGC u GCAGACAG 814 CUGUCUGC CUGAUGAG GCCGUUAGGC CGAA ICAGGAAA , 3003
1358 CCUGCUGC A GACAGUUG 815 CAACUGUC CUGAUGAG GCCGUUAGGC CGAA ICAGCAGG 3004
1362 CUGCAGAC A GUUGAGCU 816 AGCUCAAC CUGAUGAG GCCGUUAGGC CGAA IUCUGCAG 3005
1370 AGUUGAGC U GGGGUCCU 817 AGGACCCC CUGAUGAG GCCGUUAGGC CGAA ICUCAACU 3006'
1377 CUGGGGUC C UGGGUUGG 818 CCAACCCA CUGAUGAG GCCGUUAGGC CGAA IACCCCAG 3007
1378 UGGGGUCC U GGGUUGGG 819 CCCAACCC CUGAUGAG GCCGUUAGGC CGAA IGACCCCA 3008
1395j AUGGUGAC A UUUGACAG 820 CUGUCAAA CUGAUGAG GCCGUUAGGC CGAA IUCACCAU 3009
1402 CAUUUGAC A GUGCUGCC 821 GGCAGCAC CUGAUGAG GCCGUUAGGC CGAA IUCAAAUG 3010
1407 GACAGUGC U GCCCAUGU 822 ACAUGGGC CUGAUGAG GCCGUUAGGC CGAA ICACUGUC 3011
1410_| AGUGCUGC C CAUGUACA 823 UGUACAUG CUGAUGAG GCCGUUAGGC CGAA ICAGCACU 3012
1411 GUGCUGCC C AUGUACAA 824 UUGUACAU CUGAUGAG GCCGUUAGGC CGAA IGCAGCAC 3013
1412 UGCUGCCC A UGUACAAA 825 UUUGUACA CUGAUGAG GCCGUUAGGC CGAA IGGCAGCA 3014
1418 CCAUGUAC A AAGUGAAC 826 GUUCACUU CUGAUGAG GCCGUUAGGC CGAA IUACAUGG 3015
1427 AAGUGAAC U CAUACAGA 827 UCUGUAUG CUGAUGAG GCCGUUAGGC CGAA IUUCACUϋ 3016
1429 GUGAACUC A UACAGAUA 828 UAUCUGUA CUGAUGAG GCCGUUAGGC CGAA IAGUUCAC 3017
1433 ACUCAUAC A GAUAAACA 829 UGUUUAUC CUGAUGAG GCCGUUAGGC CGAA IUAUGAGU 3018
1441 AGAUAAAC A GUGGCAGU 830 ACUGCCAC CUGAUGAG GCCGUUAGGC CGAA lUUUAUCU 3019
1447 ACAGUGGC A GUGACAGG 831 CCUGUCAC CUGAUGAG GCCGUUAGGC CGAA ICCACUGU 3020
1453 GCAGUGAC A GGGACACA 832 UGUGUCCC CUGAUGAG GCCGUUAGGC CGAA IUCACUGC 3021
1459 ACAGGGAC A CACUCGCC 833 GGCGAGUG CUGAUGAG GCCGUUAGGC CGAA IUCCCUGU 3022
1461 AGGGACAC A CUCGCCAA 834 UUGGCGAG CUGAUGAG GCCGUUAGGC CGAA lUGUCCCU 3023
1463 GGACACAC U CGCCAAAA 835 UUUUGGCG CUGAUGAG GCCGUUAGGC CGAA IUGUGUCC 3024
1467 ACACUCGC C AAAAGAUU 836 AAUCUUUU CUGAUGAG GCCGUUAGGC CGAA ICGAGUGU 3025
1468 CACUCGCC A AAAGAUUA 837 UAAUCUUU CUGAUGAG GCCGUUAGGC CGAA IGCGAGUG 3026
1478 AAGAUUAC C UGCAGCAG 838 CUGCUGCA CUGAUGAG GCCGUUAGGC CGAA IUAAUCUU 3027
1479 AGAUUACC U GCAGCAGC 839 GCUGCUGC CUGAUGAG GCCGUUAGGC CGAA IGUAAUCU 3028
1482 UUACCUGC A GCAGCUUC 840 GAAGCUGC CUGAUGAG GCCGUUAGGC CGAA ICAGGUAA 3029
1485 CCUGCAGC A GCUUCAGG 841 CCUGAAGC CUGAUGAG GCCGUUAGGC CGAA ICUGCAGG 3030
1488 GCAGCAGC U UCAGGAGG 842 CCUCCUGA CUGAUGAG GCCGUUAGGC CGAA ICUGCUGC 3031
1491 GCAGCUUC A GGAGGGAC 843 GUCCCUCC CUGAUGAG GCCGUUAGGC CGAA IAAGCUGC 3032
I
1503 GGGACGUC C AUCUGCAG 844 CUGCAGAU CUGAUGAG GCCGUUAGGC CGAA IACGUCCC 3033
1504 GGACGUCC A UCUGCAGC 845 GCUGCAGA CUGAUGAG GCCGUUAGGC CGAA IGACGUCC 3034
1507 CGUCCAUC U GCAGCGGG 846 CCCGCUGC CUGAUGAG GCCGUUAGGC CGAA IAUGGACG 3035
1510 CCAUCUGC A GCGGGCUU 847 AAGCCCGC CUGAUGAG GCCGUUAGGC CGAA ICAGAUGG 3036
1517 CAGCGGGC U UCGAUCGG 848 CCGAUCGA CUGAUGAG GCCGUUAGGC CGAA ICCCGCUG 3037
1527 CGAUCGGC A UUUACUGU 849 ACAGUAAA CUGAUGAG GCCGUUAGGC CGAA ICCGAUCG 3038
1533" GCAUUUAC U GUGAUUAG 850 CUAAUCAC CUGAUGAG GCCGUUAGGC CGAA IUAAAUGC 3039
1553 GAAAUAUC C AACUGAUG 851 CAUCAGUU CUGAUGAG GCCGUUAGGC CGAA IAUAUUUC 3040
1554 AAAUAUCC A ACUGAUGG 852 CCAUCAGU CUGAUGAG GCCGUUAGGC CGAA IGAUAUUU 3041
1557 UAUCCAAC U GAUGGAUC 853 GAUCCAUC CUGAUGAG GCCGUUAGGC CGAA IUUGGAUA 3042
1566 GAUGGAUC U GAAAUUGU 854 ACAAUUUC CUGAUGAG GCCGUUAGGC CGAA IAUCCAUC 3043
1577 AAUUGUGC U GCUGACGG 855 CCGUCAGC CUGAUGAG GCCGUUAGGC CGAA ICACAAUU 3044
1580 UGUGCUGC U GACGGAUG 856 CAUCCGUC CUGAUGAG GCCGUUAGGC CGAA ICAGCACA 3045
1597 GGGAAGAC A ACACUAUA 857 UAUAGUGU CUGAUGAG GCCGUUAGGC CGAA IUCUUCCC 3046
1600 AAGACAAC A CUAUAAGU^ 858 ACUUAUAG CUGAUGAG GCCGUUAGGC CGAA IUUGUCUϋ 3047
1602 GACAACAC U AUAAGUGG 859 CCACUUAU CUGAUGAG GCCGUUAGGC CGAA IUGUUGUC 3048
1615 GUGGGUGC U UUAACGAG 860 CUCGUUAA CUGAUGAG GCCGUUAGGC CGAA ICACCCAC 3049
1627 ACGAGGUC A AACAAAGU 861 ACUUUGUU CUGAUGAG GCCGUUAGGC CGAA IACCUCGU 3050
1631 GGUCAAAC A AAGUGGUG 862 CACCACUU CUGAUGAG GCCGUUAGGC CGAA IUUUGACC 3051
1641 AGUGGUGC C AUCAUCCA 863 UGGAUGAU CUGAUGAG GCCGUUAGGC CGAA ICACCACU 3052
1642 GUGGUGCC A UCAUCCAC 864 GUGGAUGA CUGAUGAG GCCGUUAGGC CGAA IGCACCAC 3053
1645 GUGCCAUC A UCCACACA 8&5 UGUGUGGA CUGAUGAG GCCGUUAGGC CGA IAUGGCAC 3054
1648 CCAUCAUC C ACACAGUC 866 GACUGUGU CUGAUGAG GCCGUUAGGC CGTAA. IAUGAUGG 3055
1649 CAUCAUCC A CACAGUCG 867 CGACUGUG CUGAUGAG GCCGUUAGGC CGAA IGAUGAUG 3056
1651 UCAUCCAC A CAGUCGCU 868 AGCGACUG CUGAUGAG GCCGUUAGGC CGAA lUGGAUGA 3057
1653 AUCCACAC A GUCGCUUU 869 AAAGCGAC CUGAUGAG GCCGUUAGGC CGAA IUGUGGAU 3058
1659 ACAGUCGC U UUGGGGCC 870 GGCCCCAA CUGAUGAG GCCGUUAGGC CGAA ICGACUGU 3059
1667 UUUGGGGC C CUCUGCAG 871 CUGCAGAG CUGAUGAG GCCGUUAGGC CGAA ICCCCAAA 3060
1668 UUGGGGCC C UCUGCAGC 872 GCUGCAGA CUGAUGAG GCCGUUAGGC CGAA IGCCCCAA 3061
1669 UGGGGCCC U CUGCAGCU 873 AGCUGCAG CUGAUGAG GCCGUUAGGC CGAA IGGCCCCA 3062
1671 GGGCCCUC u GCAGCUCA 874 UGAGCUGC CUGAUGAG GCCGUUAGGC CGAA IAGGGCCC 3063
1674 CCCUCUGC A GCUCAAGA 875 UCUUGAGC CUGAUGAG GCCGUUAGGC CGAA ICAGAGGG 3064
1677 UCUGCAGC U CAAGAACU 876 AGUUCUUG CUGAUGAG GCCGUUAGGC CGAA ICUGCAGA 3065
1679 UGCAGCUC A AGAACUAG 877 CUAGUUCU CUGAUGAG GCCGUUAGGC CGAA IAGCUGCA 3066
1685 UCAAGAAC U AGAGGAGC 878 GCUCCUCU CUGAUGAG GCCGUUAGGC CGAA IUUCUUGA 3067
1694 AGAGGAGC U GUCCAAAA 879 UUUUGGAC CUGAUGAG GCCGUUAGGC CGAA ICUCCUCU 3068
1698 GAGCUGUC C AAAAUGAC 880 GUCAUUUU CUGAUGAG GCCGUUAGGC CGAA IACAGCUC 3069
1699 AGCUGUCC A AAAUGACA 881 UGUCAUUU CUGAUGAG GCCGUUAGGC CGAA IGACAGCU 3070
1707 AAAAUGAC A GGAGGUUU 882 AAACCUCC CUGAUGAG GCCGUUAGGC CGAA IUCAUUUU 3071
1718 AGGUUUAC A GACAUAUG 883 CAUAUGUC CUGAUGAG GCCGUUAGGC CGAA IUAAACCU 3072
1722 UUACAGAC A UAUGCUUC 884 GAAGCAUA CUGAUGAG GCCGUUAGGC CGAA IUCUGUAA 3073
1728 ACAUAUGC U UCAGAUCA 885 UGAUCUGA CUGAUGAG GCCGUUAGGC CGAA ICAUAUGU 3074
1731 UAUGCUUC A GAUCAAGU 886 ACUUGAUC CUGAUGAG GCCGUUAGGC CGAA IAAGCAUA 3075
1736 UUCAGAUC A AGUUCAGA 887 UCUGAACU CUGAUGAG GCCGUUAGGC CGAA IAUCUGAA _, 3076
1742 UCAAGUUC A GAACAAUG 888 CAUUGUUC CUGAUGAG GCCGUUAGGC CGAA IAACUUGA 3077
1747 UUCAGAAC A AUGGCCUC 889 GAGGCCAU CUGAUGAG GCCGUUAGGC CGAA IUUCUGAA 3078
1753 ACAAUGGC C UCAUUGAU 890 AUCAAUGA CUGAUGAG GCCGUUAGGC CGAA ICCAUUGU 3079
1754 CAAUGGCC U CAUUGAUG 891 CAUCAAUG CUGAUGAG GCCGUUAGGC CGAA IGCCAUUG 3080
1756 AUGGCCUC A UUGAUGCU 892 AGCAUCAA CUGAUGAG GCCGUUAGGC CGAA IAGGCCAU 3081
1764 AUUGAUGC U UUUGGGGC 893 GCCCCAAA CUGAUGAG GCCGUUAGGC CGAA ICAUCAAU 3082
1773 UUUGGGGC C CUUUCAUC 894 GAUGAAAG CUGAUGAG GCCGUUAGGC CGAA ICCCCAAA 3083
1774 UUGGGGCC C UUUCAUCA 895 UGAUGAAA CUGAUGAG GCCGUUAGGC CGAA IGCCCCAA 3084
1775 UGGGGCCC U UUCAUCAG 896 CUGAUGAA CUGAUGAG GCCGUUAGGC CGAA IGGCCCCA 3085
1779 GCCCUUUC A UCAGGAAA 897 UUUCCUGA CUGAUGAG GCCGUUAGGC CGAA IAAAGGGC 3086
1782 CUUUCAUC A GGAAAUGG 898 CCAUUUCC CUGAUGAG GCCGUUAGGC CGAA IAUGAAAG 3087
1794 AAUGGAGC U GUCUCUCA 899 UGAGAGAC CUGAUGAG GCCGUUAGGC CGAA ICUCCAUU 3088
1798 GAGCUGUC U CUCAGCGC 900 GCGCUGAG CUGAUGAG GCCGUUAGGC CGAA IACAGCUC 3089
1800 GCUGUCUC u CAGCGCUC 901 GAGCGCUG CUGAUGAG GCCGUUAGGC CGAA IAGACAGC 3090
1802 UGUCUCUC A GCGCUCCA 902 UGGAGCGC CUGAUGAG GCCGUUAGGC CGAA IAGAGACA 3091
1807 CUCAGCGC U CCAUCCAG 903 CUGGAUGG CUGAUGAG GCCGUUAGGC CGAA ICGCUGAG 3092
1809 CAGCGCUC C AUCCAGCU 904 AGCUGGAU CUGAUGAG GCCGUUAGGC CGAA IAGCGCUG 3093
1810 AGCGCUCC A UCCAGCUU 905 AAGCUGGA CUGAUGAG GCCGUUAGGC CGAA IGAGCGCU 3094
1813 GCUCCAUC C AGCUUGAG 906 CUCAAGCU CUGAUGAG GCCGUUAGGC CGAA IAUGGAGC 3095
1814 CUCCAUCC A GCUUGAGA 907 UCUCAAGC CUGAUGAG GCCGUUAGGC CGAA IGAUGGAG 3096
1817 CAUCCAGC U UGAGAGUA 908 UACUCUCA CUGAUGAG GCCGUUAGGC CGAA ICUGGAUG 3097
1836 GGAUUAAC C CUCCAGAA 909 UUCUGGAG CUGAUGAG GCCGUUAGGC CGAA IUUAAUCC 3098
1837 GAUUAACC C UCCAGAAC 910 GUUCUGGA CUGAUGAG GCCGUUAGGC CGAA IGUUAAUC 3099
1838 AUUAACCC U CCAGAACA 911 UGUUCUGG CUGAUGAG GCCGUUAGGC CGAA IGGUUAAU 3100
1840 UAACCCUC C AGAACAGC 912 GCUGUUCU CUGAUGAG GCCGUUAGGC CGAA IAGGGUUA 3101
1841 AACCCUCC A GAACAGCC 913 GGCUGUUC CUGAUGAG GCCGUUAGGC CGAA IGAGGGUU 3102
1846 UCCAGAAC A GCCAGUGG 914 CCACUGGC CUGAUGAG GCCGUUAGGC CGAA IUUCUGGA 3103
1849 AGAACAGC C AGUGGAUG 915 CAUCCACU CUGAUGAG GCCGUUAGGC CGAA ICUGUUCU 3104
1850 GAACAGCC A GUGGAUGA 916 UCAUCCAC CUGAUGAG GCCGUUAGGC CGAA IGCUGUUC 3105
1864 UGAAUGGC A CAGUGAUC 917 GAUCACUG CUGAUGAG GCCGUUAGGC CGAA ICCAUUCA 3106
1866 AAUGGCAC A GUGAUCGU 918 ACGAUCAC CUGAUGAG GCCGUUAGGC CGAA IUGCCAUU 3107
1879 UCGUGGAC A GCACCGUG 919 CACGGUGC CUGAUGAG GCCGUUAGGC CGAA IUCCACGA 3108
1882 UGGACAGC A CCGUGGGA 920 UCCCACGG CUGAUGAG GCCGUUAGGC CGAA ICUGUCCA 3109
1884 GACAGCAC C GUGGGAAA 921 UUUCCCAC CUGAUGAG GCCGUUAGGC CGAA IUGCUGUC 3110
1897 GAAAGGAC A CUUUGUUU 922 AAACAAAG CUGAUGAG GCCGUUAGGC CGAA IUCCUUUC 3111
1899 AAGGACAC U UUGUUUCU 923 AGAAACAA CUGAUGAG GCCGUUAGGC CGAA lUGUCCUU 3112
1907 UUUGUϋUC u UAUCACCU 924 AGGUGAUA CUGAUGAG GCCGUUAGGC CGAA IAAACAAA 3113
1912 UUCUUAUC A CCUGGACA 925 UGUCCAGG CUGAUGAG GCCGUUAGGC CGAA IAUAAGAA 3114
1914 CUUAUCAC C UGGACAAC 926 GUUGUCCA CUGAUGAG GCCGUUAGGC CGAA IUGAUAAG 3115
1915 UUAUCACC U GGACAACG 927 CGUUGUCC CUGAUGAG GCCGUUAGGC CGAA IGUGAUAA 3116
1920 ACCUGGAC A ACGCAGCC 928 GGCUGCGU CUGAUGAG GCCGUUAGGC CGAA IUCCAGGU 3117
1925 GACAACGC A GCCUCCCC 929 GGGGAGGC CUGAUGAG GCCGUUAGGC CGAA ICGUUGUC 3118
1928 AACGCAGC C UCCCCAAA 930 UUUGGGGA CUGAUGAG GCCGUUAGGC CGAA ICUGCGUU 3119
1929 ACGCAGCC U CCCCAAAU 931 AUUUGGGG CUGAUGAG GCCGUUAGGC CGAA IGCUGCGU 3120
1931 GCAGCCUC C CCAAAUCC 932 GGAUUUGG CUGAUGAG GCCGUUAGGC CGAA IAGGCUGC 3121
1932 CAGCCUCC C CAAAUCCU 933 AGGAUUUG CUGAUGAG GCCGUUAGGC CGAA IGAGGCUG 3122
1933 AGCCUCCC C AAAUCCUU 934 AAGGAUUU CUGAUGAG GCCGUUAGGC CGAA IGGAGGCU 3123
1934 GCCUCCCC A AAUCCUUC 935 GAAGGAUU CUGAUGAG GCCGUUAGGC CGAA IGGGAGGC 3124
1939 CCCAAAUC C UUCUCUGG 936 CCAGAGAA CUGAUGAG GCCGUUAGGC CGAA IAUUUGGG 3125
1940 CCAAAUCC U UCUCUGGG 937 CCCAGAGA CUGAUGAG GCCGUUAGGC CGAA IGAUUUGG 3126
1943 AAUCCUUC U CUGGGAUC 938 GAUCCCAG CUGAUGAG GCCGUUAGGC CGAA IAAGGAUU 3127
1945 UCCUUCUC u GGGAUCCC 939 GGGAUCCC CUGAUGAG GCCGUUAGGC CGAA IAGAAGGA 3128
1952 CUGGGAUC c CAGUGGAC 940 GUCCACUG CUGAUGAG GCCGUUAGGC CGAA IAUCCCAG 3129
1953 UGGGAUCC c AGUGGACA 941 UGUCCACU CUGAUGAG GCCGUUAGGC CGAA IGAUCCCA 3130
1954 GGGAUCCC A GUGGACAG 942 CUGUCCAC CUGAUGAG GCCGUUAGGC CGAA IGGAUCCC 3131
1961 CAGUGGAC A GAAGCAAG 943 CUUGCUUC CUGAUGAG GCCGUUAGGC CGAA IUCCACUG 3132
1967 ACAGAAGC A AGGUGGCU 944 AGCCACCU CUGAUGAG GCCGUUAGGC CGAA ICUUCUGU 3133
197 ,—5 AAGGUGGC U UUGUAGUG 945 CACUACAA CUGAUGAG GCCGUUAGGC CGAA ICCACCUU 3134
1987 UAGUGGAC A AAAACACC 946 GGUGUUUU CUGAUGAG GCCGUUAGGC CGAA IUCCACUA 3135
1993 ACAAAAAC A CCAAAAUG 947 CAUUUUGG CUGAUGAG GCCGUUAGGC CGAA IUUUUUGU 3136
1995 AAAAACAC C AAAAUGGC 948 GCCAUUUU CUGAUGAG GCCGUUAGGC CGAA IUGUUUUU 3137
1996 AAAACACC A AAAUGGCC 949 GGCCAUUU CUGAUGAG GCCGUUAGGC CGAA IGUGUUUU 3138
2004 AAAAUGGC C UACCUCCA 950 UGGAGGUA CUGAUGAG GCCGUUAGGC CGAA ICCAUUUU 3139
2005 AAAUGGCC u ACCUCCAA 951 UUGGAGGU CUGAUGAG GCCGUUAGGC CGAA IGCCAUUU 3140
2008 UGGCCUAC c UCCAAAUC 952 GAUUUGGA CUGAUGAG GCCGUUAGGC CGAA IUAGGCCA 3141
2009 GGCCUACC u CCAAAUCC 953 GGAUUUGG CUGAUGAG GCCGUUAGGC CGAA IGUAGGCC 3142
2011 CCUACCUC c AAAUCCCA 954 UGGGAUUU CUGAUGAG GCCGUUAGGC CGAA IAGGUAGG 3143
2012 CUACCUCC A AAUCCCAG 955 CUGGGAUU CUGAUGAG GCCGUUAGGC CGAA IGAGGUAG 3144
2017 UCCAAAUC C CAGGCAUU 956 AAUGCCUG CUGAUGAG GCCGUUAGGC CGAA lAUUUGGA 3145
2018 CCAAAUCC C AGGCAUUG 957 CAAUGCCU CUGAUGAG GCCGUUAGGC CGAA IGAUUUGG 3146
2019 CAAAUCCC A GGCAUUGC 958 GCAAUGCC CUGAUGAG GCCGUUAGGC CGAA IGGAUUUG 3147
2023 UCCCAGGC A UUGCUAAG 959 CUUAGCAA CUGAUGAG GCCGUUAGGC CGAA ICCUGGGA 3148
2028 GGCAUUGC U AAGGUUGG 960 CCAACCUU CUGAUGAG GCCGUUAGGC CGAA ICAAUGCC 3149
2038 AGGUUGGC A CUUGGAAA 961 UUUCCAAG CUGAUGAG GCCGUUAGGC CGAA ICCAACCU 3150
2040 GUUGGCAC U UGGAAAUA 962 UAUUUCCA CUGAUGAG GCCGUUAGGC CGAA IUGCCAAC 3151
2050 GGAAAUAC A GUCUGCAA 963 UUGCAGAC CUGAUGAG GCCGUUAGGC CGAA IUAUUUCC 3152
2054 AUACAGUC U GCAAGCAA 964 UUGCUUGC CUGAUGAG GCCGUUAGGC CGAA IACUGUAU 3153
2057 CAGUCUGC A AGCAAGCU 965 AGCUUGCU CUGAUGAG GCCGUUAGGC CGAA ICAGACUG 3154
2061 CUGCAAGC A AGCUCACA 966 UGUGAGCU CUGAUGAG GCCGUUAGGC CGAA ICUUGCAG 3155
2065 AAGCAAGC U CACAAACC 967 GGUUUGUG CUGAUGAG GCCGUUAGGC CGAA ICUUGCUU 3156
2067 GCAAGCUC A CAAACCUU 968 AAGGUUUG CUGAUGAG GCCGUUAGGC CGAA IAGCUUGC 3157
2069 AAGCUCAC A AACCUUGA 969 UCAAGGUU CUGAUGAG GCCGUUAGGC CGAA IUGAGCUU 3158
2073 UCACAAAC C UUGACCCU 970 AGGGUCAA CUGAUGAG GCCGUUAGGC CGAA lUUUGUGA 3159
2074 CACAAACC u UGACCCUG 971 CAGGGUCA CUGAUGAG GCCGUUAGGC CGAA IGUUUGUG 3160
2079 ACCUUGAC C CUGACUGU 972 ACAGUCAG CUGAUGAG GCCGUUAGGC CGAA lUCAAGGU 3161
2080 CCUUGACC c UGACUGUC 973 GACAGUCA CUGAUGAG GCCGUUAGGC CGAA IGUCAAGG 3162
2081 CUUGACCC u GACUGUCA 974 UGACAGUC CUGAUGAG GCCGUUAGGC CGAA IGGUCAAG 3163
2085 ACCCUGAC u GUCACGUC 975 GACGUGAC CUGAUGAG GCCGUUAGGC CGAA IUCAGGGU 3164
2089 UGACUGUC A CGUCCCGU 976 ACGGGACG CUGAUGAG GCCGUUAGGC CGAA lACAGUCA 3165
2094 GUCACGUC c CGUGCGUC 977 GACGCACG CUGAUGAG GCCGUUAGGC CGAA IACGUGAC 3166
2095 UCACGUCC c GUGCGUCC 978 GGACGCAC CUGAUGAG GCCGUUAGGC CGAA IGACGUGA 3167
2103 CGUGCGUC c AAUGCUAC 979 GUAGCAUU CUGAUGAG GCCGUUAGGC CGAA IACGCACG 3168
2104 GUGCGUCC A AUGCUACC 980 GGUAGCAU CUGAUGAG GCCGUUAGGC CGAA IGACGCAC 3169
2109 UCCAAUGC U ACCCUGCC 981 GGCAGGGU CUGAUGAG GCCGUUAGGC CGAA ICAUUGGA 3170
2112 AAUGCUAC C CUGCCUCC 982 GGAGGCAG CUGAUGAG GCCGUUAGGC CGAA IUAGCAUU 3171
2113 AUGCUACC C UGCCUCCA 983 UGGAGGCA CUGAUGAG GCCGUUAGGC CGAA IGUAGCAU 3172
2114 UGCUACCC U GCCUCCAA 984 UUGGAGGC CUGAUGAG GCCGUUAGGC CGAA IGGUAGCA 3173
2117 UACCCUGC C UCCAAUUA 985 UAAUUGGA CUGAUGAG GCCGUUAGGC CGAA ICAGGGUA 3174
2118 ACCCUGCC U CCAAUUAC 986 GUAAUUGG CUGAUGAG GCCGUUAGGC CGAA IGCAGGGU 3175
2120 CCUGCCUC c AAUUACAG 987 CUGUAAUU CUGAUGAG GCCGUUAGGC CGAA IAGGCAGG 3176
2121 CUGCCUCC A AUUACAGU 988 ACUGUAAU CUGAUGAG GCCGUUAGGC CGAA IGAGGCAG 3177
2127 CCAAUUAC A GUGACUUC 989 GAAGUCAC CUGAUGAG GCCGUUAGGC CGAA IUAAUUGG 3178
2133 ACAGUGAC U UCCAAAAC 990 GUUUUGGA CUGAUGAG GCCGUUAGGC CGAA IUCACUGU 3179
2136 GUGACUUC C AAAACGAA 991 UUCGUUUU CUGAUGAG GCCGUUAGGC CGAA IAAGUCAC 3180
2137 UGACUUCC A AAACGAAC 992 GUUCGUUU CUGAUGAG GCCGUUAGGC CGAA IGAAGUCA 3181
2146 AAACGAAC A AGGACACC 993 GGUGUCCU CUGAUGAG GCCGUUAGGC CGAA IUUCGUUU 3182
2152 ACAAGGAC A CCAGCAAA 994 UUUGCUGG CUGAUGAG GCCGUUAGGC CGAA IUCCUUGU 3183
2154 AAGGACAC C AGCAAAUU 995 AAUUUGCU CUGAUGAG GCCGUUAGGC CGAA lUGUCCUU 3184
2155 AGGACACC A GCAAAUUC 996 GAAUUUGC CUGAUGAG GCCGUUAGGC CGAA IGUGUCCU 3185
2158 ACACCAGC A AAUUCCCC 997 GGGGAAUU CUGAUGAG GCCGUUAGGC CGAA ICUGGUGU 3186
2164 GCAAAUUC C CCAGCCCU 998 AGGGCUGG CUGAUGAG GCCGUUAGGC CGAA IAAUUUGC 3187
2165 CAAAUUCC C CAGCCCUC 999 GAGGGCUG CUGAUGAG GCCGUUAGGC CGAA IGAAUUUG 3188
2166 AAAUUCCC C AGCCCUCU 1000 AGAGGGCU CUGAUGAG GCCGUUAGGC CGAA IGGAAUUU 3189
2167 AAUUCCCC A GCCCUCUG 1001 CAGAGGGC CUGAUGAG GCCGUUAGGC CGAA IGGGAAUU 3190
2170 UCCCCAGC C CUCUGGUA 1002 UACCAGAG CUGAUGAG GCCGUUAGGC CGAA ICUGGGGA 3191
2171 CCCCAGCC C UCUGGUAG 1003 CUACCAGA CUGAUGAG GCCGUUAGGC CGAA IGCUGGGG 3192
2172 CCCAGCCC U CUGGUAGU 1004 ACUACCAG CUGAUGAG GCCGUUAGGC CGAA IGGCUGGG 3193
2174 CAGCCCUC u GGUAGUUU 1005 AAACUACC CUGAUGAG GCCGUUAGGC CGAA IAGGGCUG 3194
2187 GUUUAUGC A AAUAUUCG 1006 CGAAUAUU CUGAUGAG GCCGUUAGGC CGAA ICAUAAAC 3195
2197 AUAUUCGC C AAGGAGCC 1007 GGCUCCUU CUGAUGAG GCCGUUAGGC CGAA ICGAAUAU 3196
2198 UAUUCGCC A AGGAGCCU 1008 AGGCUCCU CUGAUGAG GCCGUUAGGC CGAA IGCGAAUA 3197
2205 CAAGGAGC C UCCCCAAU 1009 AUUGGGGA CUGAUGAG GCCGUUAGGC CGAA ICUCCUUG 3198
2206 AAGGAGCC U CCCCAAUU 1010 AAUUGGGG CUGAUGAG GCCGUUAGGC CGAA IGCUCCUU 3199
2208 GGAGCCUC c CCAAUUCU 1011 AGAAUUGG CUGAUGAG GCCGUUAGGC CGAA IAGGCUCC 3200
2209 GAGCCUCC c CAAUUCUC 1012 GAGAAUUG CUGAUGAG GCCGUUAGGC CGAA IGAGGCUC 3201
2210 AGCCUCCC c AAUUCUCA 1013 UGAGAAUU CUGAUGAG GCCGUUAGGC CGAA IGGAGGCU 3202
2211 GCCUCCCC A AUUCUCAG 1014 CUGAGAAU CUGAUGAG GCCGUUAGGC CGAA IGGGAGGC 3203
2216 CCCAAUUC U CAGGGCCA 1015 UGGCCCUG CUGAUGAG GCCGUUAGGC CGAA IAAUUGGG 3204
2218 CAAUUCUC A GGGCCAGU 1016 ACUGGCCC CUGAUGAG GCCGUUAGGC CGAA IAGAAUUG 3205
2223 CUCAGGGC C AGUGUCAC 1017 GUGACACU CUGAUGAG GCCGUUAGGC CGAA ICCCUGAG 3206
2224 UCAGGGCC A GUGUCACA 1018 UGUGACAC CUGAUGAG GCCGUUAGGC CGAA IGCCCUGA 3207
2230 CCAGUGUC A CAGCCCUG 1019 CAGGGCUG CUGAUGAG GCCGUUAGGC CGAA IACACUGG 3208
2232 AGUGUCAC A GCCCUGAU 1020 AUCAGGGC CUGAUGAG GCCGUUAGGC CGAA lUGACACU 3209
2235 GUCACAGC C CUGAUUGA 1021 UCAAUCAG CUGAUGAG GCCGUUAGGC CGAA ICUGUGAC 3210
2236 UCACAGCC C UGAUUGAA 1022 UUCAAUCA CUGAUGAG GCCGUUAGGC CGAA IGCUGUGA 3211
2237 CACAGCCC U GAUUGAAU 1023 AUUCAAUC CUGAUGAG GCCGUUAGGC CGAA IGGCUGUG 3212
2247 AUUGAAUC A GUGAAUGG^ 1024 CCAUUCAC CUGAUGAG GCCGUUAGGC CGAA IAUUCAAU 3213
2262 GGAAAAAC A GUUACCUU 1025 AAGGUAAC CUGAUGAG GCCGUUAGGC CGAA IUUUUUCC 3214
2268 ACAGUUAC C UUGGAACU 1026 AGUUCCAA CUGAUGAG GCCGUUAGGC CGAA lUAACUGU 3215
2269 CAGUUACC U UGGAACUA 1027 UAGUUCCA CUGAUGAG GCCGUUAGGC CGAA IGUAACUG 3216
2276 CUUGGAAC U ACUGGAUA 1028 UAUCCAGU CUGAUGAG GCCGUUAGGC CGAA IUUCCAAG 3217
2279 GGAACUAC u GGAUAAUG 1029 CAUUAUCC CUGAUGAG GCCGUUAGGC CGAA IUAGUUCC 3218
2292 AAUGGAGC A GGUGCUGA 1030 UCAGCACC CUGAUGAG GCCGUUAGGC CGAA ICUCCAUU 3219
2298 GCAGGUGC U GAUGCUAC 1031 GUAGCAUC CUGAUGAG GCCGUUAGGC CGAA ICACCUGC 3220
2304 GCUGAUGC U ACUAAGGA 1032 UCCUUAGU CUGAUGAG GCCGUUAGGC CGAA ICAUCAGC 3221
2307 GAUGCUAC u AAGGAUGA 1033 UCAUCCUU CUGAUGAG GCCGUUAGGC CGAA IUAGCAUC 3222
2323 ACGGUGUC u ACUCAAGG 1034 CCUUGAGU CUGAUGAG GCCGUUAGGC CGAA IACACCGU 3223
2326 GUGUCUAC u CAAGGUAU 1035 AUACCUUG CUGAUGAG GCCGUUAGGC CGAA IUAGACAC 3224
2328 GUCUACUC A AGGUAUUU 1036 AAAUACCU CUGAUGAG GCCGUUAGGC CGAA IAGUAGAC 3225
2338 GGUAUUUC A CAACUUAU 1037 AUAAGUUG CUGAUGAG GCCGUUAGGC CGAA IAAAUACC 3226
2340 UAUUUCAC A ACUUAUGA 1038 UCAUAAGU CUGAUGAG GCCGUUAGGC CGAA lUGAAAUA 3227
2343 UUCACAAC U UAUGACAC 1039 GUGUCAUA CUGAUGAG GCCGUUAGGC CGAA IUUGUGAA 3228
2350 CUUAUGAC A CGAAUGGU 1040 ACCAUUCG CUGAUGAG GCCGUUAGGC CGAA IUCAUAAG 3229
2365 GUAGAUAC A GUGUAAAA 1041 UUUUACAC CUGAUGAG GCCGUUAGGC CGAA lUAUCUAC 3230
2382 GUGCGGGC U CUGGGAGG 1042 CCUCCCAG CUGAUGAG GCCGUUAGGC CGAA ICCCGCAC 3231
2384 GCGGGCUC U GGGAGGAG 1043 CUCCUCCC CUGAUGAG GCCGUUAGGC CGAA IAGCCCGC 3232
2400 GUUAACGC A GCCAGACG 1044 CGUCUGGC CUGAUGAG GCCGUUAGGC CGAA ICGUUAAC 3233
2403 AACGCAGC C AGACGGAG 1045 CUCCGUCU CUGAUGAG GCCGUUAGGC CGAA ICUGCGUU 3234
2404 ACGCAGCC A GACGGAGA 1046 UCUCCGUC CUGAUGAG GCCGUUAGGC CGAA IGCUGCGU 3235
2420 AGUGAUAC C CCAGCAGA 1047 UCUGCUGG CUGAUGAG GCCGUUAGGC CGAA lUAUCACU 3236
2421 GUGAUACC C CAGCAGAG 1048 CUCUGCUG CUGAUGAG GCCGUUAGGC CGAA IGUAUCAC 3237
2422 UGAUACCC C AGCAGAGU 1049 ACUCUGCU CUGAUGAG GCCGUUAGGC CGAA IGGUAUCA 3238
2423 GAUACCCC A GCAGAGUG 1050 CACUCUGC CUGAUGAG GCCGUUAGGC CGAA IGGGUAUC 3239
2426 ACCCCAGC A GAGUGGAG 1051 CUCCACUC CUGAUGAG GCCGUUAGGC CGAA ICUGGGGU 3240
2436 AGUGGAGC A CUGUACAU 1052 AUGUACAG CUGAUGAG GCCGUUAGGC CGAA ICUCCACU 3241
2438 UGGAGCAC U GUACAUAC 1053 GUAUGUAC CUGAUGAG GCCGUUAGGC CGAA IUGCUCCA 3242
2443 CACUGUAC A UACCUGGC 1054 GCCAGGUA CUGAUGAG GCCGUUAGGC CGAA IUACAGUG 3243
2447 GUACAUAC C UGGCUGGA 1055 UCCAGCCA CUGAUGAG GCCGUUAGGC CGAA IUAUGUAC 3244
2448 UACAUACC U GGCUGGAU 1056 AUCCAGCC CUGAUGAG GCCGUUAGGC CGAA IGUAUGUA 3245
2452 UACCUGGC U GGAUUGAG 1057 CUCAAUCC CUGAUGAG GCCGUUAGGC CGAA ICCAGGUA 3246
2474 UGAAAUAC A AUGGAAUC 1058 GAUUCCAU CUGAUGAG GCCGUUAGGC CGAA IUAUUUCA 3247
2483 AUGGAAUC C ACCAAGAC 1059 GUCUUGGU CUGAUGAG GCCGUUAGGC CGAA IAUUCCAU 3248
2484 UGGAAUCC A CCAAGACC 1060 GGUCUUGG CUGAUGAG GCCGUUAGGC CGAA IGAUUCCA 3249
2486 GAAUCCAC C AAGACCUG 1061 CAGGUCUU CUGAUGAG GCCGUUAGGC CGAA IUGGAUUC 3250
2487 AAUCCACC A AGACCUGA 1062 UCAGGUCU CUGAUGAG GCCGUUAGGC CGAA IGUGGAUU 3251
2492 ACCAAGAC C UGAAAUUA 1063 UAAUUUCA CUGAUGAG GCCGUUAGGC CGAA IUCUUGGU 3252
2493 CCAAGACC U GAAAUUAA 1064 UUAAUUUC CUGAUGAG GCCGUUAGGC CGAA IGUCUUGG 3253
2516 UGAUGUUC A ACACAAGC 1065 GCUUGUGU CUGAUGAG GCCGUUAGGC CGAA IAACAUCA 3254
2519 UGUUCAAC A CAAGCAAG 1066 CUUGCUUG CUGAUGAG GCCGUUAGGC CGAA IUUGAACA 3255
2521 UUCAACAC A AGCAAGUG 1067 CACUUGCU CUGAUGAG GCCGUUAGGC CGAA IUGUUGAA 3256
2525 ACACAAGC A AGUGUGUU 1068 AACACACU CUGAUGAG GCCGUUAGGC CGAA ICUUGUGU 3257
2536 UGUGUUUC A GCAGAACA 1069 UGUUCUGC CUGAUGAG GCCGUUAGGC CGAA lAAACACA 3258
2539 GUUUCAGC A GAACAUCC 1070 GGAUGUUC CUGAUGAG GCCGUUAGGC CGAA ICUGAAAC 3259
2544 AGCAGAAC A UCCUCGGG 1071 CCCGAGGA CUGAUGAG GCCGUUAGGC CGAA IUUCUGCU 3260
2547 AGAACAUC C UCGGGAGG 1072 CCUCCCGA CUGAUGAG GCCGUUAGGC CGAA IAUGUUCU 3261
2548 GAACAUCC U CGGGAGGC 1073 GCCUCCCG CUGAUGAG GCCGUUAGGC CGAA IGAUGUUC 3262
2557 CGGGAGGC U CAUUUGUG 1074 CACAAAUG CUGAUGAG GCCGUUAGGC CGAA ICCUCCCG 3263
2559 GGAGGCUC A UUUGUGGC 1075 GCCACAAA CUGAUGAG GCCGUUAGGC CGAA IAGCCUCC 3264
2568 UUUGUGGC U UCUGAUGU 1076 ACAUCAGA CUGAUGAG GCCGUUAGGC CGAA ICCACAAA 3265
2571 GUGGCUUC U GAUGUCCC 1077 GGGACAUC CUGAUGAG GCCGUUAGGC CGAA IAAGCCAC 3266
2578 CUGAUGUC C CAAAUGCU 1078 AGCAUUUG CUGAUGAG GCCGUUAGGC CGAA IACAUCAG 3267
2579 UGAUGUCC C AAAUGCUC 1079 GAGCAUUU CUGAUGAG GCCGUUAGGC CGAA IGACAUCA 3268
2580 GAUGUCCC A AAUGCUCC 1080 GGAGCAUU CUGAUGAG GCCGUUAGGC CGAA IGGACAUC 3269
2586 CCAAAUGC U CCCAUACC 1081 GGUAUGGG CUGAUGAG GCCGUUAGGC CGAA ICAUUUGG 3270
2588 AAAUGCUC C CAUACCUG 1082 CAGGUAUG CUGAUGAG GCCGUUAGGC CGAA IAGCAUUU 3271
2589 AAUGCUCC C AUACCUGA 1083 UCAGGUAU CUGAUGAG GCCGUUAGGC CGAA IGAGCAUU 3272
2590 AUGCUCCC A UACCUGAU 1084 AUCAGGUA CUGAUGAG GCCGUUAGGC CGAA IGGAGCAU 3273
2594 UCCCAUAC C UGAUCUCU 1085 AGAGAUCA CUGAUGAG GCCGUUAGGC CGAA IUAUGGGA 3274
2595 CCCAUACC U GAUCUCUU 1086 AAGAGAUC CUGAUGAG GCCGUUAGGC CGAA IGUAUGGG 3275
2600 ACCUGAUC u CUUCCCAC 1087 GUGGGAAG CUGAUGAG GCCGUUAGGC CGAA IAUCAGGU 3276
2602 CUGAUCUC u UCCCACCU 1088 AGGUGGGA CUGAUGAG GCCGUUAGGC CGAA IAGAUCAG 3277
2605 AUCUCUUC c CACCUGGC 1089 GCCAGGUG CUGAUGAG GCCGUUAGGC CGAA IAAGAGAU 3278
2606 UCUCUUCC c ACCUGGCC 1090 GGCCAGGU CUGAUGAG GCCGUUAGGC CGAA IGAAGAGA 3279
2607 cucuuccc A CCUGGCCA 1091 UGGCCAGG CUGAUGAG GCCGUUAGGC CGAA IGGAAGAG 3280
2609 CUUCCCAC C UGGCCAAA 1092 UUUGGCCA CUGAUGAG GCCGUUAGGC CGAA IUGGGAAG 3281
2610 UUCCCACC U GGCCAAAU 1093 AUUUGGCC CUGAUGAG GCCGUUAGGC CGAA IGUGGGAA 3282
2614 CACCUGGC c AAAUCACC 1094 GGUGAUUU CUGAUGAG GCCGUUAGGC CGAA ICCAGGUG 3283
2615 ACCUGGCC A AAUCACCG 1095 CGGUGAUU CUGAUGAG GCCGUUAGGC CGAA IGCCAGGU 3284
2620 GCCAAAUC A CCGACCUG 1096 CAGGUCGG CUGAUGAG GCCGUUAGGC CGAA IAUUUGGC 3285
2622 CAAAUCAC C GACCUGAA 1097 UUCAGGUC CUGAUGAG GCCGUUAGGC CGAA IUGAUUUG 3286
2626 UCACCGAC c UGAAGGCG 1098 CGCCUUCA CUGAUGAG GCCGUUAGGC CGAA IUCGGUGA 3287
2627 CACCGACC u GAAGGCGG 1099 CCGCCUUC CUGAUGAG GCCGUUAGGC CGAA IGUCGGUG 3288
2642 GGAAAUUC A CGGGGGCA 1100 UGCCCCCG CUGAUGAG GCCGUUAGGC CGAA IAAUUUCC 3289
2650 ACGGGGGC A GUCUCAUU 1101 AAUGAGAC CUGAUGAG GCCGUUAGGC CGAA ICCCCCGU 3290
2654 GGGCAGUC U CAUUAAUC 1102 GAUUAAUG CUGAUGAG GCCGUUAGGC CGAA IACUGCCC 3291
2656 GCAGUCUC A UUAAUCUG 1103 CAGAUUAA CUGAUGAG GCCGUUAGGC CGAA IAGACUGC 3292
2663 CAUUAAUC U GACUUGGA 1104 UCCAAGUC CUGAUGAG GCCGUUAGGC CGAA IAUUAAUG 3293
2667 AAUCUGAC U UGGACAGC 1105 GCUGUCCA CUGAUGAG GCCGUUAGGC CGAA IUCAGAUU 3294
2673 ACUUGGAC A GCUCCUGG 1106 CCAGGAGC CUGAUGAG GCCGUUAGGC CGAA IUCCAAGU 3295
2676 UGGACAGC U CCUGGGGA 1107 UCCCCAGG CUGAUGAG GCCGUUAGGC CGAA ICUGUCCA 3296
2678 GACAGCUC C UGGGGAUG 1108 CAUCCCCA CUGAUGAG GCCGUUAGGC CGAA IAGCUGUC 3297
2679 ACAGCUCC U GGGGAUGA 1109 UCAUCCCC CUGAUGAG, GCCGUUAGGC CGAA IGAGCUGU 3298
2695 AUUAUGAC C AUGGAACA 1110 UGUUCCAU CUGAUGAG GCCGUUAGGC CGAA lUCAUAAU 3299
2696 UUAUGACC A UGGAACAG 1111 CUGUUCCA CUGAUGAG GCCGUUAGGC CGAA IGUCAUAA 3300
2703 CAUGGAAC A GCUCACAA 1112 UUGUGAGC CUGAUGAG GCCGUUAGGC CGAA IUUCCAUG 3301
2706 GGAACAGC U CACAAGUA 1113 UACUUGUG CUGAUGAG GCCGUUAGGC CGAA ICUGUUCC 3302
2708 AACAGCUC A CAAGUAUA 1114 UAUACUUG CUGAUGAG GCCGUUAGGC CGAA lAGCUGUU 3303
2710 CAGCUCAC A AGUAUAUC 1115 GAUAUACU CUGAUGAG GCCGUUAGGC CGAA IUGAGCUG 3304
2719 AGUAUAUC A UUCGAAUA 1116 UAUUCGAA CUGAUGAG GCCGUUAGGC CGAA IAUAUACU 3305
2733c AUAAGUAC A AGUAUUCU 1117 AGAAUACU CUGAUGAG GCCGUUAGGC CGAA IUACUUAU 3306
2741- AAGUAUUC U UGAUCUCA 1118 UGAGAUCA CUGAUGAG GCCGUUAGGC CGAA IAAUACUU 3307
2747 UCUUGAUC U CAGAGACA 1119 UGUCUCUG CUGAUGAG GCCGUUAGGC CGAA IAUCAAGA 3308
2749 UUGAUCUC A GAGACAAG 1120 CUUGUCUC CUGAUGAG GCCGUUAGGC CGAA IAGAUCAA 3309
2755 UCAGAGAC A AGUUCAAU 1121 AUUGAACU CUGAUGAG GCCGUUAGGC CGAA IUCUCUGA 3310
2761 ACAAGUUC A AUGAAUCU 1122 AGAUUCAU CUGAUGAG GCCGUUAGGC CGAA IAACUUGU 3311
2769 AAUGAAUC U CUUCAAGU 1123 ACUUGAAG CUGAUGAG GCCGUUAGGC CGAA IAUUCAUU 3312
2771 UGAAUCUC U UCAAGUGA 1124 UCACUUGA CUGAUGAG GCCGUUAGGC CGAA IAGAUUCA 3313
2774 AUCUCUUC A AGUGAAUA 1125 UAUUCACU CUGAUGAG GCCGUUAGGC CGAA IAAGAGAU 3314
2784 GUGAAUAC U ACUGCUCU 1126 AGAGCAGU CUGAUGAG GCCGUUAGGC CGAA IUAUUCAC 3315
2787 AAUACUAC U GCUCUCAU 1127 AUGAGAGC CUGAUGAG GCCGUUAGGC CGAA IUAGUAUU 3316
2790 ACUACUGC U CUCAUCCC 1128 GGGAUGAG CUGAUGAG GCCGUUAGGC CGAA ICAGUAGU 3317
2792 UACUGCUC U CAUCCCAA 1129 UUGGGAUG CUGAUGAG GCCGUUAGGC CGAA IAGCAGUA 3318
2794 CUGCUCUC A UCCCAAAG 1130 CUUUGGGA CUGAUGAG GCCGUUAGGC CGAA IAGAGCAG 3319
2797 CUCUCAUC C CAAAGGAA 1131 UUCCUUUG CUGAUGAG GCCGUUAGGC CGAA IAUGAGAG 3320
2798 UCUCAUCC C AAAGGAAG 1132 CUUCCUUU CUGAUGAG GCCGUUAGGC CGAA IGAUGAGA 3321
2799 CUCAUCCC A AAGGAAGC 1133 GCUUCCUU CUGAUGAG GCCGUUAGGC CGAA IGGAUGAG 3322
2808 AAGGAAGC C AACUCUGA 1134 UCAGAGUU CUGAUGAG GCCGUUAGGC CGAA ICUUCCUU 3323
2809 AGGAAGCC A ACUCUGAG 1135 CUCAGAGU CUGAUGAG GCCGUUAGGC CGAA IGCUUCCU 3324
2812 AAGCCAAC U CUGAGGAA 1136 UUCCUCAG CUGAUGAG GCCGUUAGGC CGAA IUUGGCUU 3325
2814 GCCAACUC U GAGGAAGU 1137 ACUUCCUC CUGAUGAG GCCGUUAGGC CGAA IAGUUGGC 3326
2824 AGGAAGUC U UUUUGUUU 1138 AAACAAAA CUGAUGAG GCCGUUAGGC CGAA IACUUCCU 3327
2837 GUUUAAAC C AGAAAACA 1139 UGUUUUCU CUGAUGAG GCCGUUAGGC CGAA IUUUAAAC 3328
2838 UUUAAACC A GAAAACAU 1140 AUGUUUUC CUGAUGAG GCCGUUAGGC CGAA IGUUUAAA 3329
2845 CAGAAAAC A UUACUUUU 1141 AAAAGUAA CUGAUGAG GCCGUUAGGC CGAA IUUUUCUG 3330
2850 AACAUUAC U UUUGAAAA 1142 UUUUCAAA CUGAUGAG GCCGUUAGGC CGAA IUAAUGUU 3331
2863 AAAAUGGC A CAGAUCUU 1143 AAGAUCUG CUGAUGAG GCCGUUAGGC CGAA ICCAUUUU 3332
2865 AAUGGCAC A GAUCUUUϋ 1144 AAAAGAUC CUGAUGAG GCCGUUAGGC CGAA lUGCCAUU 3333
2870 CACAGAUC U UUUCAUUG 1145 CAAUGAAA CUGAUGAG GCCGUUAGGC CGAA IAUCUGUG 3334
2875 AUCUUUUC A UUGCUAUU 1146 AAUAGCAA CUGAUGAG GCCGUUAGGC CGAA lAAAAGAU 3335
2880 UUCAUUGC U AUUCAGGC 1147 GCCUGAAU CUGAUGAG GCCGUUAGGC CGAA ICAAUGAA 3336
2885 UGCUAUUC A GGCUGUUG 1148 GAACAGCC CUGAUGAG GCCGUUAGGC CGAA IAAUAGCA 3337
2889 AUUCAGGC U GUUGAUAA 1149 UUAUCAAC CUGAUGAG GCCGUUAGGC CGAA ICCUGAAU 3338
2906 GGUCGAUC U GAAAUCAG 1150 CUGAUUUC CUGAUGAG GCCGUUAGGC CGAA IAUCGACC 3339
2913 CUGAAAUC A GAAAUAUC 1151 GAUAUUUC CUGAUGAG GCCGUUAGGC CGAA IAUUUCAG 3340
2922 GAAAUAUC C AACAUUGC 1152 GCAAUGUU CUGAUGAG GCCGUUAGGC CGAA IAUAUUUC 3341
2923 AAAUAUCC A ACAUUGCA 1153 UGCAAUGU CUGAUGAG GCCGUUAGGC CGAA IGAUAUUU . 3342
2926 UAUCCAAC A UUGCACGA 1154 UCGUGCAA CUGAUGAG GCCGUUAGGC CGAA IUUGGAUA 3343
2931 AACAUUGC A CGAGUAUC 1155 GAUACUCG CUGAUGAG GCCGUUAGGC CGAA ICAAUGUU 3344
2940 CGAGUAUC U UUGUUUAU 1156 AUAAACAA CUGAUGAG GCCGUUAGGC CGAA IAUACUCG 3345
2951 GUUUAUUC C UGCACAGA 1157 UCUGUGGA CUGAUGAG GCCGUUAGGC CGAA IAAUAAAC 3346
2952 UUUAUUCC U CCACAGAC 1158 GUCUGUGG CUGAUGAG GCCGUUAGGC CGAA IGAAUAAA 3347
2954 UAUUCCUC C ACAGACUC 1159 GAGUCUGU CUGAUGAG GCCGUUAGGC CGAA IAGGAAUA 3348
2955 AUUCCUCC A CAGACUCC 1160 GGAGUCUG CUGAUGAG GCCGUUAGGC CGAA IGAGGAAU 3349
2957 UCCUCCAC A GACUCCGC 1161 GCGGAGUC CUGAUGAG GCCGUUAGGC CGAA IUGGAGGA 3350
2961 CCACAGAC U CCGCCAGA 1162 UCUGGCGG CUGAUGAG GCCGUUAGGC CGAA IUCUGUGG 3351
2963 ACAGACUC C GGCAGAGA 1163 UCUCUGGC CUGAUGAG GCCGUUAGGC CGAA IAGUCUGU 3352
2966 GACUCCGC C AGAGACAC 1164 GUGUCUCU CUGAUGAG GCCGUUAGGC CGAA ICGGAGUC 3353
2967 ACUCCGCC A GAGACACC 1165 GGUGUCUC CUGAUGAG GCCGUUAGGC CGAA IGCGGAGU 3354
2973 CCAGAGAC A CCUAGUCC 1166 GGACUAGG CUGAUGAG GCCGUUAGGC CGAA IUCUCUGG 3355
2975 AGAGACAC C UAGUCCUG 1167 CAGGACUA CUGAUGAG GCCGUUAGGC CGAA IUGUCUCU 3356
2976 GAGACACC U AGUCCUGA 1168 UCAGGACU CUGAUGAG GCCGUUAGGC CGAA IGUGUCUC 3357
2981 ACCUAGUC C UGAUGAAA 1169 UUUCAUCA CUGAUGAG GCCGUUAGGC CGAA IACUAGGU 3358
2982 CCUAGUCC u GAUGAAAC 1170 GUUUCAUC CUGAUGAG GCCGUUAGGC CGAA IGACUAGG 3359
2994 GAAACGUC u GCUCCUUG 1171 CAAGGAGC CUGAUGAG GCCGUUAGGC CGAA IACGUUUC 3360
2997 ACGUCUGC u CCUUGUCC 1172 GGACAAGG CUGAUGAG GCCGUUAGGC CGAA ICAGACGU 3361
2999 GUCUGCUC C UUGUCCUA 1173 UAGGACAA CUGAUGAG GCCGUUAGGC CGAA IAGCAGAC 3362
3000 UCUGCUCC u UGUCCUAA 1174 UUAGGACA CUGAUGAG GCCGUUAGGC CGAA IGAGCAGA 3363
3005 UCCUUGUC c UAAUAUUC 1175 GAAUAUUA CUGAUGAG GCCGUUAGGC CGAA IACAAGGA 3364
3006 CCUUGUCC u AAUAUUCA 1176 UGAAUAUU CUGAUGAG GCCGUUAGGC CGAA IGACAAGG 3365
3014 UAAUAUUC A UAUCAACA 1177 UGUUGAUA CUGAUGAG GCCGUUAGGC CGAA lAAUAUUA 3366
3019 UUCAUAUC A ACAGCACC 1178 GGUGCUGU CUGAUGAG GCCGUUAGGC CGAA lAUAUGAA 3367
3022 AUAUCAAC A GCACCAUU 1179 AAUGGUGC CUGAUGAG GCCGUUAGGC CGAA lUUGAUAU 3368
3025 UCAACAGC A CCAUUCCU 1180 AGGAAUGG CUGAUGAG GCCGUUAGGC CGAA ICUGUUGA 3369
3027 AACAGCAC c AUUCCUGG 1181 CCAGGAAU CUGAUGAG GCCGUUAGGC CGAA IUGCUGUU 3370
3028 ACAGCACC A UUCCUGGC 1182 GCCAGGAA CUGAUGAG GCCGUUAGGC CGAA IGUGCUGU 3371
3032 CACCAUUC c UGGCAUUC 1183 GAAUGCCA CUGAUGAG GCCGUUAGGC CGAA IAAUGGUG 3372
3033 ACCAUUCC U GGCAUUCA 1184 UGAAUGCC CUGAUGAG GCCGUUAGGC CGAA IGAAUGGU 3373
3037 UUCCUGGC A UUCACAUU 1185 AAUGUGAA CUGAUGAG GCCGUUAGGC CGAA ICCAGGAA 3374
3041 UGGCAUUC A CAUUUUAA 1186 UUAAAAUG CUGAUGAG GCCGUUAGGC CGAA IAAUGCCA 3375
3043 GCAUUCAC A UUUUAAAA 1187 UUUUAAAA CUGAUGAG GCCGUUAGGC CGAA IUGAAUGC 3376
3077 AGGAGAAC U GCAGCUGU 1188 ACAGCUGC CUGAUGAG GCCGUUAGGC CGAA IUUCUCCU 3377
3080 AGAACUGC A GCUGUCAA 1189 UUGACAGC CUGAUGAG GCCGUUAGGC CGAA ICAGUUCU 3378
3083 ACUGCAGC U GUCAAUAG 1190 CUAUUGAC CUGAUGAG GCCGUUAGGC CGAA ICUGCAGU 3379
3087 CAGCUGUC A AUAGCCUA 1191 UAGGCUAU CUGAUGAG GCCGUUAGGC CGAA IACAGCUG 3380
3093 UCAAUAGC C UAGGGCUG 1192 CAGCCCUA CUGAUGAG GCCGUUAGGC CGAA ICUAUUGA 3381
3094 CAAUAGCC U AGGGCUGA 1193 UCAGCCCU CUGAUGAG GCCGUUAGGC CGAA IGCUAUUG 3382
3100 CCUAGGGC U GAAUUUUU 1194 AAAAAUUC CUGAUGAG GCCGUUAGGC CGAA ICCCUAGG 3383
3112 UUUUUGUC A GAUAAAUA 1195 UAUUUAUC CUGAUGAG GCCGUUAGGC CGAA IACAAAAA 3384
3130 AAUAAAUC A UUCAUCCU 1196 AGGAUGAA CUGAUGAG GCCGUUAGGC CGAA IAUUUAUU 3385
3134 AAUCAUUC A UCCUUUUU 1197 AAAAAGGA CUGAUGAG GCCGUUAGGC CGAA lAAUGAUU 3386
3137 CAUUCAUC C UUUUUUUG 1198 CAAAAAAA CUGAUGAG GCCGUUAGGC CGAA IAUGAAUG 3387
3138 AUUCAUCC U UUUUUUGA 1199 UCAAAAAA CUGAUGAG GCCGUUAGGC CGAA IGAUGAAU 3388
3160 AAAUUUUC U AAAAUGUA 1200 UACAUUUU CUGAUGAG GCCGUUAGGC CGAA IAAAAUUϋ 3389
3177 UUUUAGAC U UCCUGUAG 1201 CUACAGGA CUGAUGAG GCCGUUAGGC CGAA IUCUAAAA 3390
3267 UUUUAGAC U UCCUGUAG 1201 CUACAGGA CUGAUGAG GCCGUUAGGC CGAA IUCUAAAA 3390
3180 UAGACUUC C UGUAGGGG 1202 CCCCUACA CUGAUGAG GCCGUUAGGC CGAA IAAGUCUA 3391
3270 UAGACUUC C UGUAGGGG 1202 CCCCUACA CUGAUGAG GCCGUUAGGC CGAA IAAGUCUA 3391
3181 AGACUUCC U GUAGGGGG 1203 CCCCCUAC CUGAUGAG GCCGUUAGGC CGAA IGAAGUCU 3392
3271 AGACUUCC u GUAGGGGG 1203 CCCCCUAC CUGAUGAG GCCGUUAGGC CGAA IGAAGUCU 3392
3198 CGAUAUAC u AAAUGUAU 1204 AUACAUUU CUGAUGAG GCCGUUAGGC CGAA lUAUAUCG 3393
3251 CGAUAUAC u AAAUGUAU 1204 AUACAUUU CUGAUGAG GCCGUUAGGC CGAA lUAUAUCG 3393
3214 UAUAGUAC A UUUAUACU 1205 AGUAUAAA CUGAUGAG GCCGUUAGGC CGAA lUACUAUA 3394
3222 AUUUAUAC U AAAUGUAU 1206 AUACAUUU CUGAUGAG GCCGUUAGGC CGAA lUAUAAAU 3395
Figure imgf000085_0001
Input Sequence = NM_001285. Cut Site = CH/.
Arm Length = 8. Core Sequence = CUGAUGAG GCCGUUAGGC CGAA
Underlined region can be any X sequence or linker, as described herein.
NM_001285 (Homo sapiens chloride channel, calcium activated, 1 (CLCAl) mRNA, 3311 bp)
249.021
Table V: Human CLCAl G-cleaver Ribozyme and Target Sequence
Pos Substrate Seq ID Ribozyme RzSeq
No.' ID No.
40 AUAUAAUU G AAUAUUUU 1211 AAAAUAUU UGAUG GCAUGCACUAUGC GCG AAUUAUAU 3400
67 GGGAGCAU G AAGAGGUG 1212 CACCUCUU UGAUG GCAUGCACUAUGC GCG AUGCUCCC 3401
78 GAGGUGUU G AGGUUAUG 1213 CAUAACCϋ UGAUG GCAUGCACUAUGC GCG AACACCUC ' 3402
106 GCACAGCU G AAGGCAGA 1214 UCUGCCUU UGAUG GCAUGCACUAUGC GCG AGCUGUGC 3403
134 ACAAGUAC G CAAUUUGA 1215 UCAAAUUG UGAUG GCAUGCACUAUGC GCG GUACUUGU ' 3404
141 CGCAAUUU G AGACUAAG 1216 CUUAGUCU UGAUG GCAUGCACUAUGC GCG AAAUUGCG 3405
172 CUCCUAUU G AAGACAAG 1217 CUUGUCUU UGAUG GCAUGCACUAUGC GCG AAUAGGAG 3406
223 AGACCUGU G AUAAACCA 1218 UGGUUUAU UGAUG GCAUGCACUAUGC GCG ACAGGUCU 3407
237 CCACUUCC G AUAAGUUG 1219 CAACUUAU UGAUG GCAUGCACUAUGC GCG GGAAGUGG 3408
312 CGUAACCC G CAUUUUCC 1220 GGAAAAUG UGAUG GCAUGCACUAUGC GCG GGGUUACG 3409
384 UUCAUCUU G AUUCUUCA 1221 UGAAGAAU UGAUG GCAUGCACUAUGC GCG AAGAUGAA 3410
411 GGGGCCCU G AGUAAUUC 1222 GAAUUACU UGAUG GCAUGCACUAUGC GCG AGGGCCCC 3411
432 AUUCAGCU G AACAACAA 1223 UUGUUGUU UGAUG GCAUGCACUAUGC GCG AGCUGAAU 3412
448 AUGGCUAU G AAGGCAUU 1224 AAUGCCUU UGAUG GCAUGCACUAUGC GCG AUAGCCAU 3413
463 UUGUCGUU G CAAUCGAC 1225 GUCGAUUG UGAUG GCAUGCACUAUGC GCG AACGACAA 3414
469 UUGCAAUC G ACCCCAAU 1226 AUUGGGGU UGAUG GCAUGCACUAUGC GCG GAUUGCAA ' 3415
480 CCCAAUGU G CCAGAAGA 1227 UCUUCUGG UGAUG GCAUGCACUAUGC GCG ACAUUGGG 3416
490 CAGAAGAU G AAACACUC 1228 GAGUGUUϋ UGAUG GCAUGCACUAUGC GCG AUCUUCUG 3417
522 GACAUGGU G ACCCAGGC 1229 GCCUGGGU UGAUG GCAUGCACUAUGC GCG ACCAUGUC 3418
547 AUGUGUUU G AAGCUACA 1230 UGUAGCUU UGAUG GCAUGCACUAUGC GCG AAACAGAU 3419
563 AGGAAAGC G AUUUUAUU 1231 AAUAAAAU UGAUG GCAUGCACUAUGC GCG GCUUUCCU 3420
583 AAAAUGUU G CCAUUUUG 1232 CAAAAUGG UGAUG GCAUGCACUAUGC GCG AACAUUUU 3421
591 GCCAUUUU G AUUCCUGA 1233 UCAGGAAU UGAUG GCAUGCACUAUGC GCG AAAAUGGC 3422
598 UGAUUCCU G AAACAUGG 1234 CCAUGUUU UGAUG GCAUGCACUAUGC GCG AGGAAUCA 3423
619 CAAAGGCU G ACUAUGUG 1235 CACAUAGU UGAUG GCAUGCACUAUGC GCG AGCCUUUG 3424
627 GACUAUGU G AGACCAAA 1236 UUUGGUCU UGAUG GCAUGCACUAUGC GCG ACAUAGUC 3425
640 CAAAACUU G AGACCUAC 1237 GUAGGUCU UGAUG GCAUGCACUAUGC GCG AAGUUUUG 3426
655 ACAAAAAU G CUGAUGUU 1238 AACAUCAG UGAUG GCAUGCACUAUGC GCG AUUUUUGU 3427
658 AAAAUGCU G AUGUUCUG 1239 CAGAACAU UGAUG GCAUGCACUAUGC GCG AGCAUUUU 3428
670 UUCUGGUU G CUGAGUCU 1240 AGACUCAG UGAUG GCAUGCACUAUGC GCG AACCAGAA 3429
673 UGGUUGCU G AGUCUACU 1241 AGUAGACU UGAUG GCAUGCACUAUGC GCG AGCAACCA 3430
694 CAGGUAAU G AUGAACCC 1242 GGGUUCAU UGAUG GCAUGCACUAUGC GCG AUUACCUG 3431
697 GUAAUGAU G AACCCUAC 1243 GUAGGGUU UGAUG GCAUGCACUAUGC GCG AUCAUUAC 3432
709 CCUACACU G AGCAGAUG 1244 CAUCUGCU UGAUG GCAUGCACUAUGC GCG AGUGUAGG 3433
739 AGAAGGGU G AAAGGAUC 1245 GAUCCUUU UGAUG GCAUGCACUAUGC GCG ACCCUUCU 3434
760 UCACUCCU G AUUUCAUU 1246 AAUGAAAU UGAUG GCAUGCACUAUGC GCG AGGAGUGA 3435
769 AUUUCAUU G CAGGAAAA 1247 UUUUCCUG UGAUG GCAUGCACUAUGC GCG AAUGAAAU 3436
787 AGUUAGCU G AAUAUGGA 1248 UCCAUAUϋ UGAUG GCAUGCACUAUGC GCG AGCUAACU ~1 3437
820 UUGUCCAU G AGUGGGCU 1249 AGCCCACU UGAUG GCAUGCACUAUGC GCG AUGGACAA 3438
836 UCAUCUAC G AUGGGGAG 1250 CUCCCCAU UGAUG GCAUGCACUAUGC GCG GUAGAUGA 3439
850 GAGUAUUU G ACGAGUAC 1251 GUACUCGU UGAUG GCAUGCACUAUGC GCG AAAUACUC 3440
853 UAUUUGAC G AGUACAAU 1252 AUUGUACU UGAUG GCAUGCACUAUGC GCG GUCAAAUA 3441
865 ACAAUAAU G AUGAGAAA 1253 UUUCUCAU UGAUG GCAUGCACUAUGC GCG AUUAUUGU 3442
868 AUAAUGAU G AGAAAUUC 1254 GAAUUUCU UGAUG GCAUGCACUAUGC GCG AUCAUUAU 3443
980 CAAAAGAU G CACAUUCA 1255 UGAAUGUG UGAUG GCAUGCACUAUGC GCG AUCUUUUG 3444
1009 GACUCUAU G AAAAAGGA 1256 UCCUUϋUU UGAUG GCAUGCACUAUGC GCG AUAGAGUC 3445
1021 AAGGAUGU G AGUUUGUU 1257 AACAAACU UGAUG GCAUGCACUAUGC GCG ACAUCCUU 3446
1040 CCAAUCCC G CCAGACGG 1258 CCGUCUGG UGAUG GCAUGCACUAUGC GCG GGGAUUGG 3447
1069 UAAUGUUU G CACAACAU 1259 AUGUUGUG UGAUG GCAUGCACUAUGC GCG AAACAUUA 3448
1081 AACAUGUU G AUUCUAUA 1260 UAUAGAAU UGAUG GCAUGCACUAUGC GCG AACAUGUU 3449
1093 CUAUAGUϋ G AAUUCUGU 1261 ACAGAAUU UGAUG GCAUGCACUAUGC GCG AACUAUAG 3450
1151 UCAAAAAU G CAAUCUCC 1262 GGAGAUUG UGAUG GCAUGCACUAUGC GCG AUUUUUGA 3451
1160 CAAUCUCC G AAGCACAU 1263 AUGUGCUU UGAUG GCAUGCACUAUGC GCG GGAGAUUG 3452
1176 UGGGAAGU G AUCCGUGA 1264 UCACGGAU UGAUG GCAUGCACUAUGC GCG ACUUCCCA 3453
1183 UGAUCCGU G AUUCUGAG 1265 CUCAGAAU UGAUG GCAUGCACUAUGC GCG ACGGAUCA 3454
1189 GUGAUUCU G AGGACUUU 1266 AAAGUCCU UGAUG GCAUGCACUAUGC GCG AGAAUCAC 3455
1215 ACUCCUAU G ACAACACA 1267 UGUGUUGU UGAUG GCAUGCACUAUGC GCG AUAGGAGU 3456
1248 UUCUCAUU G CUGCAGAU 1268 AUCUGCAG UGAUG GCAUGCACUAUGC GCG AAUGAGAA 3457
1251 UCAUUGCU G CAGAUUGG 1269 CCAAUCUG UGAUG GCAUGCACUAUGC GCG AGCAAUGA 3458
1285 UAGUCCUU G ACAAAUCU 1270 AGAUUUGU UGAUG GCAUGCACUAUGC GCG AAGGACUA 3459
1305 AGCAUGGC G ACUGGUAA 1271 UUACCAGU UGAUG GCAUGCACUAUGC GCG GCCAUGCU 3460
1316 UGGUAACC G CCUCAAUC 1272 GAUUGAGG UGAUG GCAUGCACUAUGC GCG GGUUACCA 3461
1325 CCUCAAUC G ACUGAAUC 1273 GAUUCAGU UGAUG GCAUGCACUAUGC GCG GAUUGAGG 3462
1329 AAUCGACU G AAUCAAGC 1274 GCUUGAUU UGAUG GCAUGCACUAUGC GCG AGUCGAUU 3463
1353 CUUUUCCU G CUGCAGAC 1275 GUCUGCAG UGAUG GCAUGCACUAUGC GCG AGGAAAAG 3464
1356 UUCCUGCU G CAGACAGU 1276 ACUGUCUG UGAUG GCAUGCACUAUGC GCG AGCAGGAA 3465
1366 AGACAGUU G AGCUGGGG 1277 CCCCAGCU UGAUG GCAUGCACUAUGC GCG AACUGUCU 3466
1392 GGGAUGGU G ACAUUUGA 1278 UCAAAUGU UGAUG GCAUGCACUAUGC GCG ACCAUCCC 3467
1399 UGACAUUU G ACAGUGCU 1279 AGCACUGU UGAUG GCAUGCACUAUGC GCG AAAUGUCA 3468
1405 UUGACAGU G CUGCCCAU 1280 AUGGGCAG UGAUG GCAUGCACUAUGC GCG ACUGUCAA 3469
1408 ACAGUGCU G CCCAUGUA 1281 UACAUGGG UGAUG GCAUGCACUAUGC GCG AGCACUGU 3470
1423 UACAAAGU G AACUCAUA 1282 UAUGAGUU UGAUG GCAUGCACUAUGC GCG ACUUUGUA 3471
1450 GUGGCAGU G ACAGGGAC 1283 GUCCCUGU UGAUG GCAUGCACUAUGC GCG ACUGCCAC 3472
1465 ACACACUC G CCAAAAGA 1284 UCUUUUGG UGAUG GCAUGCACUAUGC GCG GAGUGUGϋ 3473
1480 GAUUACCU G CAGCAGCU 1285 AGCUGCUG UGAUG GCAUGCACUAUGC GCG AGGUAAUC 3474
1508 GUCCAUCU G CAGCGGGC 1286 GCCCGCUG UGAUG GCAUGCACUAUGC GCG AGAUGGAC 3475
1520 CGGGCUUC G AUCGGCAU 1287 AUGCCGAU UGAUG GCAUGCACUAUGC GCG GAAGCCCG 3476
1536 UUUACUGU G AUUAGGAA 1288 UUCCUAAU UGAUG GCAUGCACUAUGC GCG ACAGUAAA 3477
1558 AUCCAACU G AUGGAUCU 1289 AGAUCCAU UGAUG GCAUGCACUAUGC GCG AGUUGGAU 3478
1567 AUGGAUCU G AAAUUGUG 1290 CACAAUUU UGAUG GCAUGCACUAUGC GCG AGAUCCAU 3479
1575 GAAAUUGU G CUGCUGAC 1291 GUCAGCAG UGAUG GCAUGCACUAUGC GCG ACAAUUUC 3480
1578 AUUGUGCU G CUGACGGA 1292 UCCGUCAG UGAUG GCAUGCACUAUGC GCG AGCACAAU 3481
1581 GUGCUGCU G ACGGAUGG 1293 CCAUCCGU UGAUG GCAUGCACUAUGC GCG AGCAGCAC 3482
1613 AAGUGGGU G CUUUAACG 1294 CGUUAAAG UGAUG GCAUGCACUAUGC GCG ACCCACUU 3483
1621 GCUUUAAC G AGGUCAAA 1295 UUUGACCU UGAUG GCAUGCACUAUGC GCG GUUAAAGC 3484
1639 AAAGUGGU G CCAUCAUC 1296 GAUGAUGG UGAUG GCAUGCACUAUGC GCG ACCACUUU 3485
1657 ACACAGUC G CUUUGGGG 1297 CCCCAAAG UGAUG GCAUGCACUAUGC GCG GACUGUGU 3486
1672 GGCCCUCU G CAGCUCAA 1298 UUGAGCUG UGAUG GCAUGCACUAUGC GCG AGAGGGCC 3487
1704 UCCAAAAU G ACAGGAGG 1299 CCUCCUGU UGAUG GCAUGCACUAUGC GCG AUUUUGGA 3488
1726 AGACAUAU G CUUCAGAU 1300 AUCUGAAG UGAUG GCAUGCACUAUGC GCG AUAUGUCU 3489
1759 GCCUCAUU G AUGCUUUU 1301 AAAAGCAU UGAUG GCAUGCACUAUGC GCG AAUGAGGC 3490
1762 UCAUUGAU G CUUUUGGG 1302 CCCAAAAG UGAUG GCAUGCACUAUGC GCG AUCAAUGA 3491
1805 CUCUCAGC G CUCCAUCC 1303 GGAUGGAG UGAUG GCAUGCACUAUGC GCG GCUGAGAG 3492
1819 UCCAGCUU G AGAGUAAG 1304 CUUACUCU UGAUG GCAUGCACUAUGC GCG AAGCUGGA 3493
1857 CAGUGGAU G AAUGGCAC 1305 GUGCCAUU UGAUG GCAUGCACUAUGC GCG AUCCACUG 3494
1869 GGCACAGU G AUCGUGGA 1306 UCCACGAU UGAUG GCAUGCACUAUGC GCG ACUGUGCC 3495
1923 UGGACAAC G CAGCCUCC 1307 GGAGGCUG UGAUG GCAUGCACUAUGC GCG GUUGUCCA 3496
2026 CAGGCAUU G CUAAGGUU 1308 AACCUUAG UGAUG GCAUGCACUAUGC GCG AAUGCCUG 3497
2055 UACAGUCU G CAAGCAAG 1309 CUUGCUUG UGAUG GCAUGCACUAUGC GCG AGACUGUA 3498
2076 CAAACCUU G ACCCUGAC 1310 GUCAGGGU UGAUG GCAUGCACUAUGC GCG AAGGUUUG 3499
2082 UUGACCCU G ACUGUCAC 1311 GUGACAGU UGAUG GCAUGCACUAUGC GCG AGGGUCAA 3500
2098 CGUCCCGU G CGUCCAAU 1312 AUUGGACG UGAUG GCAUGCACUAUGC GCG ACGGGACG 3501
2107 CGUCCAAU G CUACCCUG 1313 CAGGGUAG UGAUG GCAUGCACUAUGC GCG AUUGGACG 3502
2115 GCUACCCU G CGUCCAAU 1314 AUUGGAGG UGAUG GCAUGCACUAUGC GCG AGGGUAGC 3503
2130 AUUACAGU G ACUUCCAA 1315 - UUGGAAGU UGAUG GCAUGCACUAUGC GCG ACUGUAAU 3504
2142 UCCAAAAC G AACAAGGA 1316 UCCUUGUU UGAUG GCAUGCACUAUGC GCG GUUUUGGA 3505
2185 UAGUUUAU G CAAAUAUU 1317 AAUAUUUG UGAUG GCAUGCACUAUGC GCG AUAAACUA 3506
2195 AAAUAUUC G CCAAGGAG 1318 CUCCUUGG UGAUG GCAUGCACUAUGC GCG GAAUAUUU 3507
2238 ACAGCCCU G AUUGAAUC 1319 GAUUCAAU UGAUG GCAUGCACUAUGC GCG AGGGCUGU 3508
2242 CCCUGAUU G AAUCAGUG 1320 CACUGAUU UGAUG GCAUGCACUAUGC GCG AAUCAGGG 3509
2250 GAAUCAGU G AAUGGAAA 1321 UUUCCAUU UGAUG GCAUGCACUAUGC GCG ACUGAUUC 3510
2296 GAGCAGGU G CUGAUGCU 1322 AGCAUCAG UGAUG GCAUGCACUAUGC GCG ACCUGCUC 3511
2299 CAGGUGCU G AUGCUACU 1323 AGUAGCAU UGAUG GCAUGCACUAUGC GCG AGCACCUG 3512
2302 GUGCUGAU G CUACUAAG 1324 CUUAGUAG UGAUG GCAUGCACUAUGC GCG AUCAGCAC 3513
2314 CUAAGGAU G ACGGUGUC 1325 GACACCGU UGAUG GCAUGCACUAUGC GCG AUCCUUAG 3514
2347 CAACUUAU G ACACGAAU 1326 AUUCGUGU UGAUG GCAUGCACUAUGC GCG AUAAGUUG 3515
2352 UAUGACAC G AAUGGUAG 1327 CUACCAUU UGAUG GCAUGCACUAUGC GCG GUGUCAUA 3516
2376 GUAAAAGU G CGGGCUCU 1328 AGAGCCCG UGAUG GCAUGCACUAUGC GCG ACUUUUAC 3517
2398 GAGUUAAC G CAGCCAGA 1329 UCUGGCUG UGAUG GCAUGCACUAUGC GCG GUUAACUC 3518
2415 CGGAGAGU G AUACCCCA 1330 UGGGGUAU UGAUG GCAUGCACUAUGC GCG ACUCUCCG 3519
2458 GCUGGAUU G AGAAUGAU 1331 AUCAUUCU UGAUG GCAUGCACUAUGC GCG AAUCCAGC 3520
2464 UUGAGAAU G AUGAAAUA 1332 UAUUUCAU UGAUG GCAUGCACUAUGC GCG AUUCUCAA 3521
2467 AGAAUGAU G AAAUACAA 1333 UUGUAUUU UGAUG GCAUGCACUAUGC GCG AUCAUUCU 3522
2494 CAAGACCU G AAAUUAAU 1334 AUUAAUUU UGAUG GCAUGCACUAUGC GCG AGGUCUUG 3523
2509 AUAAGGAU G AUGUUCAA 1335 UUGAACAU UGAUG GCAUGCACUAUGC GCG AUCCUUAU 3524
2572 UGGCUUCU G AUGUCCCA 1336 UGGGACAU UGAUG GCAUGCACUAUGC GCG AGAAGCCA 3525
2584 UCCCAAAU G CUCCCAUA 1337 UAUGGGAG UGAUG GCAUGCACUAUGC GCG AUUUGGGA 3526
2596 CCAUACCU G AUCUCUUC 1338 GAAGAGAU UGAUG GCAUGCACUAUGC GCG AGGUAUGG 3527
2623 AAAUCACC G ACCUGAAG 1339 CUUCAGGU UGAUG GCAUGCACUAUGC GCG GGUGAUUU 3528
2628 ACCGACCU G AAGGCGGA 1340 UCCGCCUU UGAUG GCAUGCACUAUGC GCG AGGUCGGU 3529
2664 AUUAAUCU G ACUUGGAC 1341 GUCCAAGU UGAUG GCAUGCACUAUGC GCG AGAUUAAU 3530
2686 CUGGGGAU G AUUAUGAC 1342 GUCAUAAU UGAUG GCAUGCACUAUGC GCG AUCCCCAG 3531
2692 AUGAUUAU G ACCAUGGA 1343 UCCAUGGU UGAUG GCAUGCACUAUGC GCG AUAAUCAU 3532
2723 UAUCAUUC G AAUAAGUA 1344 UACUUAUU UGAUG GCAUGCACUAUGC GCG GAAUGAUA 3533
2743 GUAUUCUU G AUCUCAGA 1345 UCUGAGAU UGAUG GCAUGCACUAUGC GCG AAGAAUAC 3534
2764 AGUUCAAU G AAUCUCUU 1346 AAGAGAUU UGAUG GCAUGCACUAUGC GCG AUUGAACU 3535
2778 CUUCAAGU G AAUACUAC 1347 GUAGUAUU UGAUG GCAUGCACUAUGC GCG ACUUGAAG 3536
2788 AUACUACU G CUCUCAUC 1348 GAUGAGAG UGAUG GCAUGCACUAUGC GCG AGUAGUAU 3537
2815 CCAACUCU G AGGAAGUC 1349 GACUUCCU UGAUG GCAUGCACUAUGC GCG AGAGUUGG 3538
2854 TJUACUUUU G AAAAUGGC 1350 GCCAUUUU UGAUG GCAUGCACUAUGC GCG AAAAGUAA 3539
2878 UUUUCAUU G CUAUUCAG 1351 CUGAAUAG UGAUG GCAUGCACUAUGC GCG AAUGAAAA 3540
2893 AGGCUGUU G AUAAGGUC 1352 GACCUUAU UGAUG GCAUGCACUAUGC GCG AACAGCCU 3541
2902 AUAAGGUC G AUCUGAAA 1353 UUUCAGAU UGAUG GCAUGCACUAUGC GCG GACCUUAU 3542
2907 GUCGAUCU G AAAUCAGA 1354 UCUGAUUU UGAUG GCAUGCACUAUGC GCG AGAUCGAC 3543
2929 CCAACAUU G CACGAGUA 1355 UACUCGUG UGAUG GCAUGCACUAUGC GCG AAUGUUGG 3544
2933 CAUUGCAC G AGUAUCUU 1356 AAGAUACU UGAUG GCAUGCACUAUGC GCG GUGCAAUG 3545
2964 CAGACUCC G CCAGAGAC 1357 GUCUCUGG UGAUG GCAUGCACUAUGC GCG GGAGUCUG 3546
2983 CUAGUCCU G AUGAAACG 1358 CGUUUCAU UGAUG GCAUGCACUAUGC GCG AGGACUAG 3547
2986 GUCCUGAU G AAACGUCU 1359 AGACGUUU UGAUG GCAUGCACUAUGC GCG AUCAGGAC 3548
2995 AAACGUCU G CUCCUUGU 1360 ACAAGGAG UGAUG GCAUGCACUAUGC GCG AGACGUUU 3549
3078 GGAGAACU G CAGCUGUC 1361 GACAGCUG UGAUG GCAUGCACUAUGC GCG AGUUCUCC 3550
3101 CUAGGGCU G AAUUUUUG 1362 CAAAAAUU UGAUG GCAUGCACUAUGC GCG AGCCCUAG 3551
3145 CUUUUUUU G AUUAUAAA 1363 UUUAUAAU UGAUG GCAUGCACUAUGC GCG AAAAAAAG 3552
3191 UAGGGGGC G AUAUACUA 1364 UAGUAUAU UGAUG GCAUGCACUAUGC GCG GCCCCCUA 3553
3244 UAGGGGGC G AUAUACUA 1364 UAGUAUAU UGAUG GCAUGCACUAUGC GCG GCCCCCUA 3553
3281 UAGGGGGC G AUAAAAUA 1365 UAUUUUAU UGAUG GCAUGCACUAUGC GCG GCCCCCUA 3554
3294 AAUAAAAU G CUAAACAA 1366 UUGUUUAG UGAUG GCAUGCACUAUGC GCG AUUUUAUU 3555
27 AAAUGGAU G UGGAAUAU 1367 AUAUUCCA UGAUG GCAUGCACUAUGC GCG AUCCAUUU 3556
52 AUUUUCUU G UUUAAGGG 1368 CCCUUAAA UGAUG GCAUGCACUAUGC GCG AAGAAAAU 3557
75 GAAGAGGU G UUGAGGUU 1369 AACCUCAA UGAUG GCAUGCACUAUGC GCG ACCUCUUC 3558
86 GAGGUUAU G UCAAGCAU 1370 AUGCUUGA UGAUG GCAUGCACUAUGC GCG AUAACCUC 3559
155 AAGAUAUU G UUAUCAUU 1371 AAUGAUAA UGAUG GCAUGCACUAUGC GCG AAUAUCUU 3560
221 AAAGACCU G UGAUAAAC 1372 GUUUAUCA UGAUG GCAUGCACUAUGC GCG AGGUCUUU 3561
253 GGAAACGU G UGUCUAUA 1373 UAUAGACA UGAUG GCAUGCACUAUGC GCG ACGUUUCC 3562
255 AAACGUGU G UCUAUAUU 1374 AAUAUAGA UGAUG GCAUGCACUAUGC GCG ACACGUUU 3563
273 UCAUAUCU G UAUAUAUA 1375 UAUAUAUA UGAUG GCAUGCACUAUGC GCG AGAUAUGA 3564
344 AGGGAGAU G UACAGCAA 1376 UUGCUGUA UGAUG GCAUGCACUAUGC GCG AUCUCCCU 3565
373 AGAGUUCU G UGUUCAUC 1377 GAUGAACA UGAUG GCAUGCACUAUGC GCG AGAACUCU 3566
375 AGUUCUGU G UUCAUCUϋ 1378 AAGAUGAA UGAUG GCAUGCACUAUGC GCG ACAGAACU 3567
457 AAGGCAUU G UCGUUGCA 1379 UGCAACGA UGAUG GCAUGCACUAUGC GCG AAUGCCUU 3568
478 ACCCCAAU G UGCCAGAA 1380 UUCUGGCA UGAUG GCAUGCACUAUGC GCG AUUGGGGU 3569
537 GCAUCUCU G UAUCUGUU 1381 AACAGAUA UGAUG GCAUGCACUAUGC GCG AGAGAUGC 3570
543 CUGUAUCU G UUUGAAGC 1382 GCUUCAAA UGAUG GCAUGCACUAUGC GCG AGAUACAG 3571
580 UCAAAAAU G UUGCCAUU 1383 AAUGGCAA UGAUG GCAUGCACUAUGC GCG AUUUUUGA 3572
625 CUGACUAU G UGAGACCA 1384 UGGUCUCA UGAUG GCAUGCACUAUGC GCG AUAGUCAG 3573
661 AUGCUGAU G UUCUGGUU 1385 AACCAGAA UGAUG GCAUGCACUAUGC GCG AUCAGCAU 3574
725 GGGCAACU G UGGAGAGA 1386 UCUCUCCA UGAUG GCAUGCACUAUGC GCG AGUUGCCC 3575
814 AGGCAUUU G UCCAUGAG 1387 CUCAUGGA UGAUG GCAUGCACUAUGC GCG AAAUGCCU 3576
911 AGUAAGAU G UUCAGCAG 1388 CUGCUGAA UGAUG GCAUGCACUAUGC GCG AUCUUACU 3577
937 GUACAAAU G UAGUAAAG 1389 CUUUACUA UGAUG GCAUGCACUAUGC GCG AUUUGUAC 3578
950 AAAGAAGU G UCAGGGAG 1390 CUCCCUGA UGAUG GCAUGCACUAUGC GCG ACUUCUUU 3579
965 AGGCAGCU G UUACACCA 1391 UGGUGUAA UGAUG GCAUGCACUAUGC GCG AGCUGCCU 3580
1019 AAAAGGAU G UGAGUUUG 1392 CAAACUCA UGAUG GCAUGCACUAUGC GCG AUCCUUUU 3581
1027 GUGAGUUU G UUCUCCAA 1393 UUGGAGAA UGAUG GCAUGCACUAUGC GCG AAACUCAC 3582
1065 UCUAUAAU G UUUGCACA 1394 UGUGCAAA UGAUG GCAUGCACUAUGC GCG AUUAUAGA 3583
1078 CACAACAU G UUGAUUCU 1395 AGAAUCAA UGAUG GCAUGCACUAUGC GCG AUGUUGUG 3584
1100 UGAAUUCU G UACAGAAC 1396 GUUCUGUA UGAUG GCAUGCACUAUGC GCG AGAAUUCA 3585
1270 AAAGAAUU G UGUGUUUA 1397 UAAACACA UGAUG GCAUGCACUAUGC GCG AAUUCUUU 3586
1272 AGAAUUGU G UGUUUAGU 1398 ACUAAACA UGAUG GCAUGCACUAUGC GCG ACAAUUCU 3587
1274 AAUUGUGU G UUUAGUCC 1399 GGACUAAA UGAUG GCAUGCACUAUGC GCG ACACAAUU 3588
1414 CUGCCCAU G UACAAAGU 1400 ACUUUGUA UGAUG GCAUGCACUAUGC GCG AUGGGCAG 3589
1534 CAUUUACU G UGAUUAGG 1401 CCUAAUCA UGAUG GCAUGCACUAUGC GCG AGUAAAUG 3590
1573 CUGAAAUU G UGCUGCUG 1402 CAGCAGCA UGAUG GCAUGCACUAUGC GCG AAUUUCAG 3591
1695 GAGGAGCU G UCCAAAAU 1403 AUUUUGGA UGAUG GCAUGCACUAUGC GCG AGCUCCUC 3592
1795 AUGGAGCU G UCUCUCAG 1404 CUGAGAGA UGAUG GCAUGCACUAUGC GCG AGCUCCAU 3593
1902 GACACUUU G UUUCUUAU 1405 AUAAGAAA UGAUG GCAUGCACUAUGC GCG AAAGUGUC 3594
1978 GUGGCUUU G UAGUGGAC 1406 GUCCACUA UGAUG GCAUGCACUAUGC GCG AAAGCCAC 3595
2086 CCCUGACU G UCACGUCC 1407 GGACGUGA UGAUG GCAUGCACUAUGC GCG AGUCAGGG 3596
2227 GGGCCAGU G UCACAGCC 1408 GGCUGUGA UGAUG GCAUGCACUAUGC GCG ACUGGCCC 3597
2320 AUGACGGU G UCUACUCA 1409 UGAGUAGA UGAUG GCAUGCACUAUGC GCG ACCGUCAU 3598
2368 GAUACAGU G UAAAAGUG 1410 CACUUUUA UGAUG GCAUGCACUAUGC GCG ACUGUAUC 3599
2439 GGAGCACU G UACAUACC 1411 GGUAUGUA UGAUG GCAUGCACUAUGC GCG AGUGCUCC 3600
2512 AGGAUGAU G UUCAACAC 1412 GUGUUGAA UGAUG GCAUGCACUAUGC GCG AUCAUCCU 3601
2529 AAGCAAGU G UGUUUCAG 1413 CUGAAACA UGAUG GCAUGCACUAUGC GCG ACUUGCUU 3602
2531 GCAAGUGU G UUUCAGCA 1414 UGCUGAAA UGAUG GCAUGCACUAUGC GCG ACACUUGC 3603
2563 GCUCAUUU G UGGCUUCU 1415 AGAAGCCA UGAUG GCAUGCACUAUGC GCG AAAUGAGC 3604
2575 CUUCUGAU G UCCCAAAU 1416 AUUUGGGA UGAUG GCAUGCACUAUGC GCG AUCAGAAG 3605
2829 GUCUUUUU G UUUAAACC 1417 GGUUUAAA UGAUG GCAUGCACUAUGC GCG AAAAAGAC 3S06
2890 UUCAGGCU G UUGAUAAG 1418 CUUAUCAA UGAUG GCAUGCACUAUGC GCG AGCCUGAA 3607
2943 GUAUCUUU G UUUAUUCC 1419 GGAAUAAA UGAUG GCAUGCACUAUGC GCG AAAGAUAC 3608
3002 UGCUCCUU G UCCUAAUA 1420 UAUUAGGA UGAUG GCAUGCACUAUGC GCG AAGGAGCA 3609
3057 AAAAUUAU G UGGAAGUG 1421 CACUUCCA UGAUG GCAUGCACUAUGC GCG AUAAUUUU 3610
3084 CUGCAGCU G UCAAUAGC 1422 GCUAUUGA UGAUG GCAUGCACUAUGC GCG AGCUGCAG 3611
3109 GAAUUUUU G UCAGAUAA 1423 UUAUCUGA UGAUG GCAUGCACUAUGC GCG AAAAAUUC 3612
3166 UCUAAAAU G UAUUUUAG 1424 CUAAAAUA UGAUG GCAUGCACUAUGC GCG AUUUUAGA 3613
3182 GACUUCCU G UAGGGGGC 1425 GCCCCCUA UGAUG GCAUGCACUAUGC GCG AGGAAGUC 3614
3272 GACUUCCU G UAGGGGGC 1425 GCCCCCUA UGAUG GCAUGCACUAUGC GCG AGGAAGUC 3614
3203 UACUAAAU G UAUAUAGU 1426 ACUAUAUA UGAUG GCAUGCACUAUGC GCG AUUUAGUA 3615
3227 UACUAAAU G UAUUCCUG 1427 CAGGAAUA UGAUG GCAUGCACUAUGC GCG AUUUAGUA 3616
3235 GUAUUCCU G UAGGGGGC 1428 GCCCCCUA UGAUG GCAUGCACUAUGC GCG AGGAAUAC 3617
3256 UACUAAAU G UAUUUUAG 1429 CUAAAAUA UGAUG GCAUGCACUAUGC GCG AUUUAGUA 3618
Input Sequence = NM_001285. Cut Site = YG/M or UG/U.
Arm Length = 8. Core Sequence = UGAUG GCAUGCACUAUGC GCG
NM_001285 (Homo sapiens chloride channel, calcium activated, 1 (CLCAl) mRNA, 3311 bp)
Table VI: Human CLCAl Zinzyme and Target Sequence 249.021
Pos Substrate Seq ID Zinzyme RzSeq ID
134 ACAAGUAC G CAAUUUGA 1215 UCAAAUUG GCCGAAAGGCGAGUGAGGUCU GUACUUGU 3619
312 CGUAACCC G CAUUUUCC 1220 GGAAAAUG GCCGAAAGGCGAGUGAGGUCU GGGUUACG 3620
463 UUGUCGUU G CAAUCGAC 1225 GUCGAUUG GCCGAAAGGCGAGUGAGGUCU AACGACAA 3621
480 CCCAAUGU G CCAGAAGA 1227 UGUUCUGG GCCGAAAGGCGAGUGAGGUCU ACAUUGGG 3622
583 AAAAUGUU G CCAUUUUG 1232 CAAAAUGG GCCGAAAGGCGAGUGAGGUCU AACAUUUU 3623
655 ACAAAAAU G CUGAUGUU 1238 AACAUCAG GCCGAAAGGCGAGUGAGGUCU AUUUUUGU 3624
670 UUCUGGUU G CUGAGUCU 1240 AGACUCAG GCCGAAAGGCGAGUGAGGUCU AACCAGAA 3625
769 AUUUCAUU G CAGGAAAA 1247 UUUUCCUG GCCGAAAGGCGAGUGAGGUCU AAUGAAAU 3626
980 CAAAAGAU G CACAUUCA 1255 UGAAUGUG GCCGAAAGGCGAGUGAGGUCU AUCUUUUG 3627
1040 CCAAUCCC G CCAGACGG 1258 CCGUCUGG GCCGAAAGGCGAGUGAGGUCU GGGAUUGG 3628
1069 UAAUGUUϋ G CACAACAU 1259 AUGUUGUG GCCGAAAGGCGAGUGAGGUCU AAACAUUA 3629
1151 UCAAAAAU G CAAUCUCC 1262 GGAGAUUG GCCGAAAGGCGAGUGAGGUCU AUUUUUGA 3630
1248 UUCUCAUU G CUGCAGAU 1268 AUCUGCAG GCCGAAAGGCGAGUGAGGUCU AAUGAGAA 3631
1251 UCAUUGCU G CAGAUUGG 1269 CCAAUCUG GCCGAAAGGCGAGUGAGGUCU AGCAAUGA 3632
1316 UGGUAACC G CCUCAAUC 1272 GAUUGAGG GCCGAAAGGCGAGUGAGGUCU GGUUACCA 3633
1353 CUUUUCCU G CUGCAGAC 1275 GUCUGCAG GCCGAAAGGCGAGUGAGGUCU AGGAAAAG 3634
1356 UUCCUGCU G CAGACAGU 1276 ACUGUCUG GCCGAAAGGCGAGUGAGGUCU AGCAGGAA 3635
1405 UUGACAGU G CUGCCCAU 1280 AUGGGCAG GCCGAAAGGCGAGUGAGGUCU ACUGUCAA 3636
1408 ACAGUGCU G CCCAUGUA 1281 UACAUGGG GCCGAAAGGCGAGUGAGGUCU AGCACUGU 3637
1465 ACACACUC G CCAAAAGA 1284 UCUUUUGG GCCGAAAGGCGAGUGAGGUCU GAGUGUGU 3638
1480 GAUUACCU G CAGCAGCU 1285 AGCUGCUG GCCGAAAGGCGAGUGAGGUCU AGGUAAUC 3639
1508 GUCCAUCU G CAGCGGGC 1286 GCCCGCUG GCCGAAAGGCGAGUGAGGUCU AGAUGGAC 3640
1575 GAAAUUGU G CUGCUGAC 1291 GUCAGCAG GCCGAAAGGCGAGUGAGGUCU ACAAUUUC 3641
1578 AUUGUGCU G CUGACGGA 1292 UCCGUCAG GCCGAAAGGCGAGUGAGGUCU AGCACAAU 3642
1613 AAGUGGGU G CUUUAACG 1294 CGUUAAAG GCCGAAAGGCGAGUGAGGUCU ACCCACUU 3643
1639 AAAGUGGU G CCAUCAUC 1296 GAUGAUGG GCCGAAAGGCGAGUGAGGUCU ACCACUUU 3644
1657 ACACAGUC G CUUUGGGG 1297 CCCCAAAG GCCGAAAGGCGAGUGAGGUCU GACUGUGU 3645
1672 GGCCCUCU G CAGCUCAA 1298 UUGAGCUG GCCGAAAGGCGAGUGAGGUCU AGAGGGCC 3646
1726 AGACAUAU G CUUCAGAU 1300 AUCUGAAG GCCGAAAGGCGAGUGAGGUCU AUAUGUCU 3647
1762 UCAUUGAU G CUUUUGGG 1302 CCCAAAAG GCCGAAAGGCGAGUGAGGUCU AUCAAUGA 3648
1805 CUCUCAGC G CUCCAUCC 1303 GGAUGGAG GCCGAAAGGCGAGUGAGGUCU GCUGAGAG 3649
1923 UGGACAAC G CAGCCUCC 1307 GGAGGCUG GCCGAAAGGCGAGUGAGGUCU GUUGUCCA 3650
2026 CAGGCAUU G CUAAGGUU 1308 AACCUUAG GCCGAAAGGCGAGUGAGGUCU AAUGCCUG 3651
2055 UACAGUCU G CAAGCAAG 1309 CUUGCUUG GCCGAAAGGCGAGUGAGGUCU AGACUGUA 3652
2098 CGUCCCGU G CGUCCAAU 1312 AUUGGACG GCCGAAAGGCGAGUGAGGUCU ACGGGACG 3653
2107 CGUCCAAU G CUACCCUG 1313 CAGGGUAG GCCGAAAGGCGAGUGAGGUCU AUUGGACG 3654
2115 GCUACCCU G CCUCCAAU 1314 AUUGGAGG GCCGAAAGGCGAGUGAGGUCU AGGGUAGC 3655
2185 UAGUUUAU G CAAAUAUU 1317 AAUAUUUG GCCGAAAGGCGAGUGAGGUCU AUAAACUA 3656
2195 AAAUAUUC G CCAAGGAG 1318 CUCCUUGG GCCGAAAGGCGAGUGAGGUCU GAAUAUUU 3657
2296 GAGCAGGU G CUGAUGCU 1322 AGCAUCAG GCCGAAAGGCGAGUGAGGUCU ACCUGCUC 3658
2302 GUGCUGAU G CUACUAAG 1324 CUUAGUAG GCCGAAAGGCGAGUGAGGUCU AUCAGCAC 3659
2376 GUAAAAGU G CGGGCUCU 1328 AGAGCCCG GCCGAAAGGCGAGUGAGGUCU ACUUUUAC 3660
2398 GAGUUAAC G CAGCCAGA 1329 UCUGGCUG GCCGAAAGGCGAGUGAGGUCU GUUAACUC 3661
2584 UCCCAAAU G CUCCCAUA 1337 UAUGGGAG GCCGAAAGGCGAGUGAGGUCU AUUUGGGA 3662
2788 AUACUACU G CUCUCAUC 1348 GAUGAGAG GCCGAAAGGCGAGUGAGGUCU AGUAGUAU 3663
2878 UUUUCAUU G CUAUUCAG 1351 CUGAAUAG GCCGAAAGGCGAGUGAGGUCU AAUGAAAA 3664
2929 CCAACAUU G CACGAGUA 1355 UACUCGUG GCCGAAAGGCGAGUGAGGUCU AAUGUUGG 3665
2964 CAGACUCC G CCAGAGAC 1357 GUCUCUGG GCCGAAAGGCGAGUGAGGUCU GGAGUCUG 3666
2995 AAACGUCU G CUCCUUGU 1360 ACAAGGAG GCCGAAAGGCGAGUGAGGUCU AGACGUUU 3667
3078 GGAGAACU G CAGCUGUC 1361 GACAGCUG GCCGAAAGGCGAGUGAGGUCU AGUUCUCC 3668
3294 AAUAAAAU G CUAAACAA 1366 UUGUUUAG GCCGAAAGGCGAGUGAGGUCU AUUUUAUU 3669
27 AAAUGGAU G UGGAAUAU 1367 AUAUUCCA GCCGAAAGGCGAGUGAGGUCU AUCCAUUU 3670
52 AUUUUCUU G UUUAAGGG 1368 CCCUUAAA GCCGAAAGGCGAGUGAGGUCU AAGAAAAU 3671
75 GAAGAGGU G UUGAGGUU 1369 AACCUCAA GCCGAAAGGCGAGUGAGGUCU ACCUCUUC 3672
86 GAGGUUAU G UCAAGCAU 1370 AUGCUUGA GCCGAAAGGCGAGUGAGGUCU AUAACCUC 3673
155 AAGAUAUU G UUAUCAUϋ 1371 AAUGAUAA GCCGAAAGGCGAGUGAGGUCU AAUAUCUU 3674
221 AAAGACCU G UGAUAAAC 1372 GUUUAUCA GCCGAAAGGCGAGUGAGGUCU AGGUCUUU 3675
253 GGAAACGU G UGUCUAUA 1373 UAUAGACA GCCGAAAGGCGAGUGAGGUCU ACGUUUCC 3676
255 AAACGUGU G UCUAUAUU 1374 AAUAUAGA GCCGAAAGGCGAGUGAGGUCU ACACGUUU 3677
273 UCAUAUCU G UAUAUAUA 1375 UAUAUAUA GCCGAAAGGCGAGUGAGGUCU AGAUAUGA 3678
344 AGGGAGAU G UACAGCAA 1376 UUGCUGUA GCCGAAAGGCGAGUGAGGUCU AUCUCCCU 3679
373 AGAGUUCU G UGUUCAUC 1377 GAUGAACA GCCGAAAGGCGAGUGAGGUCU AGAACUCU 3680
375 AGUUCUGU G UUCAUCUU 1378 AAGAUGAA GCCGAAAGGCGAGUGAGGUCU ACAGAACU 3681
457 AAGGCAUU G UCGUUGCA 1379 UGCAACGA GCCGAAAGGCGAGUGAGGUCU AAUGCCUU 3682
478 ACCCCAAU G UGCCAGAA 1380 UUCUGGCA GCCGAAAGGCGAGUGAGGUCU AUUGGGGU 3683
537 GCAUCUCU G UAUCUGUU 1381 AACAGAUA GCCGAAAGGCGAGUGAGGUCU AGAGAUGC 3684
543 CUGUAUCU G UUUGAAGC 1382 GCUUCAAA GCCGAAAGGCGAGUGAGGUCU AGAUACAG 3685
580 UCAAAAAU G UUGCCAUU 1383 AAUGGCAA GCCGAAAGGCGAGUGAGGUCU AUUUUUGA 3686
625 CUGACUAU G UGAGACCA 1384 UGGUCUCA GCCGAAAGGCGAGUGAGGUCU AUAGUCAG 3687
661 AUGCUGAU G UUCUGGUU 1385 AACCAGAA GCCGAAAGGCGAGUGAGGUCU AUCAGCAU 3688
725 GGGCAACU G UGGAGAGA 1386 UCUCUCCA GCCGAAAGGCGAGUGAGGUCU AGUUGCCC 3689
814 AGGCAUUU G UCCAUGAG 1387 CUCAUGGA GCCGAAAGGCGAGUGAGGUCU AAAUGCCU 3690
911 AGUAAGAU G UUCAGCAG 1388 CUGCUGAA GCCGAAAGGCGAGUGAGGUCU AUCUUACU 3691
937 GUACAAAU G UAGUAAAG 1389 CUUUACUA GCCGAAAGGCGAGUGAGGUCU AUUUGUAC 3692
950 AAAGAAGU G UCAGGGAG 1390 CUCCCUGA GCCGAAAGGCGAGUGAGGUCU ACUUCUUU 3693
965 AGGCAGCU G UUACACCA 1391 UGGUGUAA GCCGAAAGGCGAGUGAGGUCU AGCUGCCU 3694
1019 AAAAGGAU G UGAGUUUG 1392 CAAACUCA GCCGAAAGGCGAGUGAGGUCU AUCCUUUU 3695
1027 GUGAGUUU G UUCUCCAA 1393 UUGGAGAA GCCGAAAGGCGAGUGAGGUCU AAACUCAC 3696
1065 UCUAUAAU G UUUGCACA 1394 UGUGCAAA GCCGAAAGGCGAGUGAGGUCU AUUAUAGA 3697
1078 CACAACAU G UUGAUUCU 1395 AGAAUCAA GCCGAAAGGCGAGUGAGGUCU AUGUUGUG 3698
1100 UGAAUUCU G UACAGAAC 1396 GUUCUGUA GCCGAAAGGCGAGUGAGGUCU AGAAUUCA 3699
1270 AAAGAAUU G UGUGUUUA 1397 UAAACACA GCCGAAAGGCGAGUGAGGUCU AAUUCUUU 3700
1272 AGAAUUGU G UGUUUAGU 1398 ACUAAACA GCCGAAAGGCGAGUGAGGUCU ACAAUUCU 3701
1274 AAUUGUGU G UUUAGUCC 1399 GGACUAAA GCCGAAAGGCGAGUGAGGUCU ACACAAUU 3702
1414 CUGCCCAU G UACAAAGU 1400 ACUUUGUA GCCGAAAGGCGAGUGAGGUCU AUGGGCAG 3703
1534 CAUUUACU G UGAUUAGG 1401 CCUAAUCA GCCGAAAGGCGAGUGAGGUCU AGUAAAUG 3704
1573 CUGAAAUU G UGCUGCUG 1402 CAGCAGCA GCCGAAAGGCGAGUGAGGUCU AAUUUCAG 3705
1695 GAGGAGCU G UCCAAAAU 1403 AUUUUGGA GCCGAAAGGCGAGUGAGGUCU AGCUCCUC 3706
1795 AUGGAGCU G UCUCUCAG 1404 CUGAGAGA GCCGAAAGGCGAGUGAGGUCU AGCUCCAU 3707
1902 GACACUUU G UUUCUUAU 1405 AUAAGAAA GCCGAAAGGCGAGUGAGGUCU AAAGUGUC 3708
1978 GUGGCUUU G UAGUGGAC 1406 GUCCACUA GCCGAAAGGCGAGUGAGGUCU AAAGCCAC 3709
2086 CCCUGACU G UCACGUCC 1407 GGACGUGA GCCGAAAGGCGAGUGAGGUCU AGUCAGGG 3710
2227 GGGCCAGU G UCACAGCC 1408 GGCUGUGA GCCGAAAGGCGAGUGAGGUCU ACUGGCCC 3711
2320 AUGACGGU G UCUACUCA 1409 UGAGUAGA GCCGAAAGGCGAGUGAGGUCU ACCGUCAU 3712
2368 GAUACAGU G UAAAAGUG 1410 CACUUUUA GCCGAAAGGCGAGUGAGGUCU ACUGUAUC 3713
2439 GGAGCACU G UACAUACC 1411 GGUAUGUA GCCGAAAGGCGAGUGAGGUCU AGUGCUCC 3714
2512 AGGAUGAU G UUCAACAC 1412 GUGUUGAA GCCGAAAGGCGAGUGAGGUCU AUCAUCCU 3715
2529 AAGCAAGU G UGUUUCAG 1413 CUGAAACA GCCGAAAGGCGAGUGAGGUCU ACUUGCUU 3716
2531 GCAAGUGU G UUUCAGCA 1414 UGCUGAAA GCCGAAAGGCGAGUGAGGUCU ACACUUGC 3717
2563 GCUCAUUU G UGGCUUCU 1415 AGAAGCCA GCCGAAAGGCGAGUGAGGUCU AAAUGAGC 3718
2575 CUUCUGAU G UCCCAAAU 1416 AUUUGGGA GCCGAAAGGCGAGUGAGGUCU AUCAGAAG 3719
2829 GUCUUUUU G UUUAAACC 1417 GGUUUAAA GCCGAAAGGCGAGUGAGGUCU AAAAAGAC 3720
2890 UUCAGGCU G UUGAUAAG 1418 CUUAUCAA GCCGAAAGGCGAGUGAGGUCU AGCCUGAA 3721
2943 GUAUCUUU G UUUAUUCC 1419 GGAAUAAA GCCGAAAGGCGAGUGAGGUCU AAAGAUAC 3722
3002 UGCUCCUU G UCCUAAUA 1420 UAUUAGGA GCCGAAAGGCGAGUGAGGUCU AAGGAGCA 3723
3057 AAAAUUAU G UGGAAGUG 1421 CACUUCCA GCCGAAAGGCGAGUGAGGUCU AUAAUUUU 3724
3084 CUGCAGCU G UCAAUAGC 1422 GCUAUUGA GCCGAAAGGCGAGUGAGGUCU AGCUGCAG 3725
3109 GAAUUUUϋ G UCAGAUAA 1423 UUAUCUGA GCCGAAAGGCGAGUGAGGUCU AAAAAUUC 3726
3166 UCUAAAAU G UAUUUUAG 1424 CUAAAAUA GCCGAAAGGCGAGUGAGGUCU AUUUUAGA 3727
3182 GACUUCCU G UAGGGGGC 1425 GCCCCCUA GCCGAAAGGCGAGUGAGGUCU AGGAAGUC 3728
3272 GACUUCCU G UAGGGGGC 1425 GCCCCCUA GCCGAAAGGCGAGUGAGGUCU AGGAAGUC 3728
3203 UACUAAAU G UAUAUAGU 1426 ACUAUAUA GCCGAAAGGCGAGUGAGGUCU AUUUAGUA 3729
3227 UACUAAAU G UAUUCCUG 1427 CAGGAAUA GCCGAAAGGCGAGUGAGGUCU AUUUAGUA 3730
3235 GUAUUCCU G UAGGGGGC 1428 GCCCCCUA GCCGAAAGGCGAGUGAGGUCU AGGAAUAC 3731
3256 UACUAAAU G UAUUUUAG 1429 CUAAAAUA GCCGAAAGGCGAGUGAGGUCU AUUUAGUA 3732
15 UGCUUUUG G UACAAAUG 1430 CAUUUGUA GCCGAAAGGCGAGUGAGGUCU CAAAAGCA 3733
63 UAAGGGGA G CAUGAAGA 1431 UCUUCAUG GCCGAAAGGCGAGUGAGGUCU UCCCCUUA 3734
73 AUGAAGAG G UGUUGAGG 1432 CCUCAACA GCCGAAAGGCGAGUGAGGUCU CUCUUCAU 3735
81 GUGUUGAG G UUAUGUCA 1433 UGACAUAA GCCGAAAGGCGAGUGAGGUCU CUCAACAC 3736
91 UAUGUCAA G CAUCUGGC 1434 GCCAGAUG GCCGAAAGGCGAGUGAGGUCU UUGACAUA 3737
98 AGCAUCUG G CACAGCUG 1435 CAGCUGUG GCCGAAAGGCGAGUGAGGUCU CAGAUGCU 3738
103 CUGGCACA G CUGAAGGC 1436 GCCUUCAG GCCGAAAGGCGAGUGAGGUCU UGUGCCAG 3739
110 AGCUGAAG G CAGAUGGA 1437 UCCAUCUG GCCGAAAGGCGAGUGAGGUCU CUUCAGCU 3740
130 AUUUACAA G UACGCAAU 1438 AUUGCGUA GCCGAAAGGCGAGUGAGGUCU UUGUAAAU 3741
182 AGACAAGA G CAAUAGUA 1439 UACUAUUG GCCGAAAGGCGAGUGAGGUCU UCUUGUCU 3742
188 GAGCAAUA G UAAAACAC 1440 GUGUUUUA GCCGAAAGGCGAGUGAGGUCU UAUUGCUC 3743
202 CACAUCAG G UCAGGGGG 1441 CCCCCUGA GCCGAAAGGCGAGUGAGGUCU CUGAUGUG 3744
210 GUCAGGGG G UUAAAGAC 1442 GUCUUUAA GCCGAAAGGCGAGUGAGGUCU CCCCUGAC 3745
242 UCCGAUAA G UUGGAAAC 1443 GUUUCCAA GCCGAAAGGCGAGUGAGGUCU UUAUCGGA 3746
251 UUGGAAAC G UGUGUCUA 1444 UAGACACA GCCGAAAGGCGAGUGAGGUCU GUUUCCAA 3747
287 AUAUAAUG G UAAAGAAA 1445 UUUCUUUA GCCGAAAGGCGAGUGAGGUCU CAUUAUAU 3748
305 ACACCUUC G UAACCCGC 1446 GCGGGUUA GCCGAAAGGCGAGUGAGGUCU GAAGGUGU 3749
349 GAUGUACA G CAAUGGGG 1447 CCCCAUUG GCCGAAAGGCGAGUGAGGUCU UGUACAUC 3750
357 GCAAUGGG G CCAUUUAA 1448 UUAAAUGG GCCGAAAGGCGAGUGAGGUCU CCCAUUGC 3751
368 AUUUAAGA G UUCUGUGU 1449 ACACAGAA GCCGAAAGGCGAGUGAGGUCU UCUUAAAU 3752
406 UAGAAGGG G CCCUGAGU 1450 ACUCAGGG GCCGAAAGGCGAGUGAGGUCU CCCUUCUA 3753
413 GGCCCUGA G UAAUUCAC 1451 GUGAAUUA GCCGAAAGGCGAGUGAGGUCU UCAGGGCC 3754
429 CUCAUUCA G CUGAACAA 1452 UUGUUCAG GCCGAAAGGCGAGUGAGGUCU UGAAUGAG 3755
443 CAACAAUG G CUAUGAAG 1453 CUUCAUAG GCCGAAAGGCGAGUGAGGUCU CAUUGUUG 3756
452 CUAUGAAG G CAUUGUCG 1454 CGACAAUG GCCGAAAGGCGAGUGAGGUCU CUUCAUAG 3757
460 GCAUUGUC G UUGCAAUC 1455 GAUUGCAA GCCGAAAGGCGAGUGAGGUCU GACAAUGC 3758
520 AGGACAUG G UGACCCAG 1456 CUGGGUCA GCCGAAAGGCGAGUGAGGUCU CAUGUCCU 3759
529 UGACCCAG G CAUCUCUG 1457 CAGAGAUG GCCGAAAGGCGAGUGAGGUCU CUGGGUCA 3760
550 UGUUUGAA G CUACAGGA 1458 UCCUGUAG GCCGAAAGGCGAGUGAGGUCU UUCAAACA 3761
561 ACAGGAAA G CGAUUUUA 1459 UAAAAUCG GCCGAAAGGCGAGUGAGGUCU UUUCCUGU 3762
616 AGACAAAG G CUGACUAU 1460 AUAGUCAG GCCGAAAGGCGAGUGAGGUCU CUUUGUCU 3763
667 AUGUUCUG G UUGCUGAG 1461 CUCAGCAA GCCGAAAGGCGAGUGAGGUCU CAGAACAU 3764
675 GUUGCUGA G UCUACUCC 1462 GGAGUAGA GCCGAAAGGCGAGUGAGGUCU UCAGCAAC 3765
689 UCCUCCAG G UAAUGAUG 1463 CAUCAUUA GCCGAAAGGCGAGUGAGGUCU CUGGAGGA 3766
711 UACACUGA G CAGAUGGG 1464 CCCAUCUG GCCGAAAGGCGAGUGAGGUCU UCAGUGUA 3767
719 GCAGAUGG G CAACUGUG 1465 CACAGUUG GCCGAAAGGCGAGUGAGGUCU CCAUCUGC 3768
737 AGAGAAGG G UGAAAGGA 1466 UCCUUUCA GCCGAAAGGCGAGUGAGGUCU CCUUCUCU 3769
780 GGAAAAAA G UUAGCUGA 1467 UCAGCUAA GCCGAAAGGCGAGUGAGGUCU UUUUUUCC 3770
784 AAAAGUUA G CUGAAUAU 1468 AUAUUCAG GCCGAAAGGCGAGUGAGGUCU UAACUUϋU 3771
803 ACCACAAG G UAAGGCAU 1469 AUGCCUUA GCCGAAAGGCGAGUGAGGUCU CUUGUGGU 3772
808 AAGGUAAG G CAUUUGUC 1470 GACAAAUG GCCGAAAGGCGAGUGAGGUCU CUUACCUU 3773
822 GUCCAUGA G UGGGCUCA 1471 UGAGCCCA GCCGAAAGGCGAGUGAGGUCU UCAUGGAC 3774
826 AUGAGUGG G CUCAUCUA 1472 UAGAUGAG GCCGAAAGGCGAGUGAGGUCU CCACUCAU 3775
844 GAUGGGGA G UAUUUGAC 1473 GUCAAAUA GCCGAAAGGCGAGUGAGGUCU UCCCCAUC 3776
855 UUUGACGA G UACAAUAA 1474 UUAUUGUA GCCGAAAGGCGAGUGAGGUCU UCGUCAAA 3777
901 GAAUACAA G CAGUAAGA 1475 UCUUACUG GCCGAAAGGCGAGUGAGGUCU UUGUAUUC 3778
904 UACAAGCA G UAAGAUGU 1476 ACAUCUUA GCCGAAAGGCGAGUGAGGUCU UGCUUGUA 3779
916 GAUGUUCA G CAGGUAUU 1477 AAUACCUG GCCGAAAGGCGAGUGAGGUCU UGAACAUC 3780
920 UUCAGCAG G UAUUACUG 1478 CAGUAAUA GCCGAAAGGCGAGUGAGGUCU CUGCUGAA 3781
929 UAUUACUG G UACAAAUG 1479 CAUUUGUA GCCGAAAGGCGAGUGAGGUCU CAGUAAUA 3782
940 CAAAUGUA G UAAAGAAG 1480 CUUCUUUA GCCGAAAGGCGAGUGAGGUCU UACAUUUG 3783
948 GUAAAGAA G UGUCAGGG 1481 CCCUGACA GCCGAAAGGCGAGUGAGGUCU UUCUUUAC 3784
959 UCAGGGAG G CAGCUGUU 1482 AACAGCUG GCCGAAAGGCGAGUGAGGUCU CUCCCUGA 3785
962 GGGAGGCA G CUGUUACA 1483 UGUAACAG GCCGAAAGGCGAGUGAGGUCU UGCCUCCC 3786
994 UCAAUAAA G UUACAGGA 1484 UCCUGUAA GCCGAAAGGCGAGUGAGGUCU UUUAUUGA 3787
1023 GGAUGUGA G UUUGUUCU 1485 AGAACAAA GCCGAAAGGCGAGUGAGGUCU UCACAUCC 3788
1054 CGGAGAAG G CUUCUAUA 1486 UAUAGAAG GCCGAAAGGCGAGUGAGGUCU CUUCUCCG 3789
1090 AUUCUAUA G UUGAAUUC 1487 GAAUUCAA GCCGAAAGGCGAGUGAGGUCU UAUAGAAU 3790
1126 ACAAAGAA G CUCCAAAC 1488 GUUUGGAG GCCGAAAGGCGAGUGAGGUCU UUCUUUGU 3791
1137 CCAAACAA G CAAAAUCA 1489 UGAUUUUG GCCGAAAGGCGAGUGAGGUCU UUGUUUGG 3792
1163 UCUCCGAA G CACAUGGG 1490 CCCAUGUG GCCGAAAGGCGAGUGAGGUCU UUCGGAGA 3793
1174 CAUGGGAA G UGAUCCGU 1491 ACGGAUCA GCCGAAAGGCGAGUGAGGUCU UUCCCAUG 3794
1181 AGUGAUCC G UGAUUCUG 1492 CAGAAUCA GCCGAAAGGCGAGUGAGGUCU GGAUCACU 3795
1224 ACAACACA G CCACCAAA 1493 UUUGGUGG GCCGAAAGGCGAGUGAGGUCU UGUGUUGU 3796
1279 UGUGUUUA G UCCUUGAC 1494 GUCAAGGA GCCGAAAGGCGAGUGAGGUCU UAAACACA 3797
1298 AUCUGGAA G CAUGGCGA 1495 UCGCCAUG GCCGAAAGGCGAGUGAGGUCU UUCCAGAU 3798
1303 GAAGCAUG G CGACUGGU 1496 ACCAGUCG GCCGAAAGGCGAGUGAGGUCU CAUGCUUC 3799
1310 GGCGACUG G UAACCGCC 1497 GGCGGUUA GCCGAAAGGCGAGUGAGGUCU CAGUCGCC 3800
1336 UGAAUCAA G CAGGCCAG 1498 CUGGCCUG GCCGAAAGGCGAGUGAGGUCU UUGAUUCA 3801
1340 UCAAGCAG G CCAGCUUU 1499 AAAGCUGG GCCGAAAGGCGAGUGAGGUCU CUGCUUGA 3802
1344 GCAGGCCA G CUUUUCCU 1500 AGGAAAAG GCCGAAAGGCGAGUGAGGUCU UGGCCUGC 3803
1363 UGCAGACA G UUGAGCUG 1501 CAGCUCAA GCCGAAAGGCGAGUGAGGUCU UGUCUGCA 3804
1368 ACAGUUGA G CUGGGGUC 1502 GACCCCAG GCCGAAAGGCGAGUGAGGUCU UCAACUGU 3805
1374 GAGCUGGG G UCCUGGGU 1503 ACCCAGGA GCCGAAAGGCGAGUGAGGUCU CCCAGCUC 3806
1381 GGUCCUGG G UUGGGAUG 1504 CAUCCCAA GCCGAAAGGCGAGUGAGGUCU CCAGGACC 3807
1390 UUGGGAUG G UGACAUUU 1505 AAAUGUCA GCCGAAAGGCGAGUGAGGUCU CAUCCCAA 3808
1403 AUUUGACA G UGCUGCCC 1506 GGGCAGCA GCCGAAAGGCGAGUGAGGUCU UGUCAAAU 3809
1421 UGUACAAA G UGAACUCA 1507 UGAGUUCA GCCGAAAGGCGAGUGAGGUCU UUUGUACA 3810
1442 GAUAAACA G UGGCAGUG 1508 CACUGCCA GCCGAAAGGCGAGUGAGGUCU UGUUUAUC 3811
1445 AAACAGUG G CAGUGACA 1509 UGUCACUG GCCGAAAGGCGAGUGAGGUCU CACUGUUU 3812
1448 CAGUGGCA G UGACAGGG 1510 CCCUGUCA GCCGAAAGGCGAGUGAGGUCU UGCCACUG 3813
1483 UACCUGCA G CAGCUUCA 1511 UGAAGCUG GCCGAAAGGCGAGUGAGGUCU UGCAGGUA 3814
1486 CUGCAGCA G CUUCAGGA 1512 UCCUGAAG GCCGAAAGGCGAGUGAGGUCU UGCUGCAG 3815
1500 GGAGGGAC G UCCAUCUG 1513 CAGAUGGA GCCGAAAGGCGAGUGAGGUCU GUCCCUCC 3816
1511 CAUCUGCA G CGGGCUUC 1514 GAAGCCCG GCCGAAAGGCGAGUGAGGUCU UGCAGAUG 3817
1515 UGCAGCGG G CUUCGAUC 1515 GAUCGAAG GCCGAAAGGCGAGUGAGGUCU CCGCUGCA 3818
1525 UUCGAUCG G CAUUUACU 1516 AGUAAAUG GCCGAAAGGCGAGUGAGGUCU CGAUCGAA 3819
1607 CACUAUAA G UGGGUGCU 1517 AGCACCCA GCCGAAAGGCGAGUGAGGUCU UUAUAGUG 3820
1611 AUAAGUGG G UGCUUUAA 1518 UUAAAGCA GCCGAAAGGCGAGUGAGGUCU CCACUUAU 3821
1624 UUAACGAG G UCAAACAA 1519 UUGUUUGA GCCGAAAGGCGAGUGAGGUCU CUCGUUAA 3822
1634 CAAACAAA G UGGUGCCA 1520 UGGCACCA GCCGAAAGGCGAGUGAGGUCU UUUGUUUG 3823
1637 ACAAAGUG G UGCCAUCA 1521 UGAUGGCA GCCGAAAGGCGAGUGAGGUCU CACUUUGU 3824
1654 UCCACACA G UCGCUUUG 1522 CAAAGCGA GCCGAAAGGCGAGUGAGGUCU UGUGUGGA 3825
1665 GCUUUGGG G CCCUCUGC 1523 GCAGAGGG GCCGAAAGGCGAGUGAGGUCU CCCAAAGC 3826
1675 CCUCUGCA G CUCAAGAA 1524 UUCUUGAG GCCGAAAGGCGAGUGAGGUCU UGCAGAGG 3827
1692 CUAGAGGA G CUGUCCAA 1525 UUGGACAG GCCGAAAGGCGAGUGAGGUCU UCCUCUAG 3828
1712 GACAGGAG G UUUACAGA 1526 UCUGUAAA GCCGAAAGGCGAGUGAGGUCU CUCCUGUC 3829
1738 CAGAUCAA G UUCAGAAC i527 GUUCUGAA GCCGAAAGGCGAGUGAGGUCU UUGAUCUG 3830
1751 GAACAAUG G CCUCAUUG 1528 CAAUGAGG GCCGAAAGGCGAGUGAGGUCU CAUUGUUC 3831
1771 CUUUUGGG G CCCUUUCA 1529 UGAAAGGG GCCGAAAGGCGAGUGAGGUCU CCCAAAAG 3832
1792 GAAAUGGA G CUGUCUCU 1530 AGAGACAG GCCGAAAGGCGAGUGAGGUCU UCCAUUUC 3833
1803 GUCUCUCA G CGCUCCAU 1531 AUGGAGCG GCCGAAAGGCGAGUGAGGUCU UGAGAGAC 3834
1815 UCCAUCCA G CUUGAGAG 1532 CUCUCAAG GCCGAAAGGCGAGUGAGGUCU UGGAUGGA 3835
1823 GCUUGAGA G UAAGGGAU 1533 AUCCCUUA GCCGAAAGGCGAGUGAGGUCU UCUCAAGC 3836
1847 CCAGAACA G CCAGUGGA 1534 UCCACUGG GCCGAAAGGCGAGUGAGGUCU UGUUCUGG 3837
1851 AACAGCCA G UGGAUGAA 1535 UUCAUCCA GCCGAAAGGCGAGUGAGGUCU UGGCUGUU 3838
1862 GAUGAAUG G CACAGUGA 1536 UCACUGUG GCCGAAAGGCGAGUGAGGUCU CAUUCAUC 3839
1867 AUGGCACA G UGAUCGUG 1537 CACGAUCA GCCGAAAGGCGAGUGAGGUCU UGUGCCAU 3840
1873 CAGUGAUC G UGGACAGC 1538 GCUGUCCA GCCGAAAGGCGAGUGAGGUCU GAUCACUG 3841
1880 CGUGGACA G CACCGUGG 1539 CCACGGUG GCCGAAAGGCGAGUGAGGUCU UGUCCACG 3842
1885 ACAGCACC G UGGGAAAG 1540 CUUUCCCA GCCGAAAGGCGAGUGAGGUCU GGUGCUGU 3843
1926 ACAACGCA G CCUCCCCA 1541 UGGGGAGG GCCGAAAGGCGAGUGAGGUCU UGCGUUGU 3844
1955 GGAUCCCA G UGGACAGA 1542 UCUGUCCA GCCGAAAGGCGAGUGAGGUCU UGGGAUCC 3845
1965 GGACAGAA G CAAGGUGG 1543 CCACCUUG GCCGAAAGGCGAGUGAGGUCU UUCUGUCC 3846
1970 GAAGCAAG G UGGCUUUG 1544 CAAAGCCA GCCGAAAGGCGAGUGAGGUCU CUUGCUUC 3847
1973 GCAAGGUG G CUUUGUAG 1545 CUACAAAG GCCGAAAGGCGAGUGAGGUCU CACCUUGC 3848
1981 GCUUUGUA G UGGACAAA 1546 UUUGUCCA GCCGAAAGGCGAGUGAGGUCU UACAAAGC 3849
2002 CCAAAAUG G CCUACCUC 1547 GAGGUAGG GCCGAAAGGCGAGUGAGGUCU CAUUUUGG 3850
2021 AAUCCCAG G CAUUGCUA 1548 UAGCAAUG GCCGAAAGGCGAGUGAGGUCU CUGGGAUU 3851
2032 UUGCUAAG G UUGGCACU 1549 AGUGCCAA GCCGAAAGGCGAGUGAGGUCU CUUAGCAA 3852
2036 UAAGGUUG G CACUUGGA 1550 UCCAAGUG GCCGAAAGGCGAGUGAGGUCU CAACCUUA 3853
2051 GAAAUACA G UCUGCAAG 1551 CUUGCAGA GCCGAAAGGCGAGUGAGGUCU UGUAUUUC 3854
2059 GUCUGCAA G CAAGCUCA 1552 UGAGCUUG GCCGAAAGGCGAGUGAGGUCU UUGCAGAC 3855
2063 GCAAGCAA G CUCACAAA 1553 UUUGUGAG GCCGAAAGGCGAGUGAGGUCU UUGCUUGC 3856
2091 ACUGUCAC G UCCCGUGC 1554 GCACGGGA GCCGAAAGGCGAGUGAGGUCU GUGACAGU 3857
2096 CACGUCCC G UGCGUCCA 1555 UGGACGCA GCCGAAAGGCGAGUGAGGUCU GGGACGUG 3858
2100 UCCCGUGC G UCCAAUGC 1556 GCAUUGGA GCCGAAAGGCGAGUGAGGUCU GCACGGGA 3859
2128 CAAUUACA G UGACUUCC 1557 GGAAGUCA GCCGAAAGGCGAGUGAGGUCU UGUAAUUG 3860
2156 GGACACCA G CAAAUUCC 1558 GGAAUUUG GCCGAAAGGCGAGUGAGGUCU UGGUGUCC 3861
2168 AUUCCCCA G CCCUCUGG 1559 CCAGAGGG GCCGAAAGGCGAGUGAGGUCU UGGGGAAU 3862
2176 GCCCUCUG G UAGUUUAU 1560 AUAAACUA GCCGAAAGGCGAGUGAGGUCU CAGAGGGC 3863
2179 CUCUGGUA G UUUAUGCA 1561 UGCAUAAA GCCGAAAGGCGAGUGAGGUCU UACCAGAG 3864
2203 GCCAAGGA G CCUCCCCA 1562 UGGGGAGG GCCGAAAGGCGAGUGAGGUCU UCCUUGGC 3865
2221 UUCUCAGG G CCAGUGUC 1563 GACACUGG GCCGAAAGGCGAGUGAGGUCU CCUGAGAA 3866
2225 CAGGGCCA G UGUCACAG 1564 CUGUGACA GCCGAAAGGCGAGUGAGGUCU UGGCCCUG 3867
2233 GUGUCACA G CCCUGAUU 1565 AAUCAGGG GCCGAAAGGCGAGUGAGGUCU UGUGACAC 3868
2248 UUGAAUCA G UGAAUGGA 1566 UCCAUUCA GCCGAAAGGCGAGUGAGGUCU UGAUUCAA 3869
2263 GAAAAACA G UUACCUUG 1567 CAAGGUAA GCCGAAAGGCGAGUGAGGUCU UGUUUUUC 3870
2290 AUAAUGGA G CAGGUGCU 1568 AGCACCUG GCCGAAAGGCGAGUGAGGUCU UCCAUUAU 3871
2294 UGGAGCAG G UGCUGAUG 1569 CAUCAGCA GCCGAAAGGCGAGUGAGGUCU CUGCUCCA 3872
2318 GGAUGACG G UGUCUACU 1570 AGUAGACA GCCGAAAGGCGAGUGAGGUCU CGUCAUCC 3873
2331 UACUCAAG G UAUUUCAC 1571 GUGAAAUA GCCGAAAGGCGAGUGAGGUCU CUUGAGUA 3874
2357 CACGAAUG G UAGAUACA 1572 UGUAUCUA GCCGAAAGGCGAGUGAGGUCU CAUUCGUG 3875
2366 UAGAUACA G UGUAAAAG 1573 CUUUUACA GCCGAAAGGCGAGUGAGGUCU UGUAUCUA 3876
2374 GUGUAAAA G UGCGGGCU 1574 AGCCCGCA GCCGAAAGGCGAGUGAGGUCU UUUUACAC 3877
2380 AAGUGCGG G CUCUGGGA 1575 UCCCAGAG GCCGAAAGGCGAGUGAGGUCU CCGCACUU 3878
2392 UGGGAGGA G UUAACGCA 1576 UGCGUUAA GCCGAAAGGCGAGUGAGGUCU UCCUCCCA 3879
2401 UUAACGCA G CCAGACGG 1577 CCGUCUGG GCCGAAAGGCGAGUGAGGUCU UGCGUUAA 3880
2413 GACGGAGA G UGAUACCC 1578 GGGUAUCA GCCGAAAGGCGAGUGAGGUCU UCUCCGUC 3881
2424 AUACCCCA G CAGAGUGG 1579 CCACUCUG GCCGAAAGGCGAGUGAGGUCU UGGGGUAU 3882
2429 CCAGCAGA G UGGAGCAC 1580 GUGCUCCA GCCGAAAGGCGAGUGAGGUCU UCUGCUGG 3883
2434 AGAGUGGA G CACUGUAC 1581 GUACAGUG GCCGAAAGGCGAGUGAGGUCU UCCACUCU 3884
2450 CAUACCUG G CUGGAUUG 1582 CAAUCCAG GCCGAAAGGCGAGUGAGGUCU CAGGUAUG 3885
2523 CAACACAA G CAAGUGUG 1583 CACACUUG GCCGAAAGGCGAGUGAGGUCU UUGUGUUG 3886
2527 ACAAGCAA G UGUGUUUC 1584 GAAACACA GCCGAAAGGCGAGUGAGGUCU UUGCUUGU 3887
2537 GUGUUUCA G CAGAACAU 1585 AUGUUCUG GCCGAAAGGCGAGUGAGGUCU UGAAACAC 3888
2555 CUCGGGAG G CUCAUUUG 1586 CAAAUGAG GCCGAAAGGCGAGUGAGGUCU CUCCCGAG 3889
2566 CAUUUGUG G CUUCUGAU 1587 AUCAGAAG GCCGAAAGGCGAGUGAGGUCU CACAAAUG 3890
2612 CCCACCUG G CCAAAUCA 1588 UGAUUUGG GCCGAAAGGCGAGUGAGGUCU CAGGUGGG 3891
2632 ACCUGAAG G CGGAAAUU 1589 AAUUUCCG GCCGAAAGGCGAGUGAGGUCU CUUCAGGU 3892
2648 UCACGGGG G CAGUCUCA 1590 UGAGACUG GCCGAAAGGCGAGUGAGGUCU CCCCGUGA 3893
2651 CGGGGGCA G UCUCAUUA 1591 UAAUGAGA GCCGAAAGGCGAGUGAGGUCU UGCCCCCG 3894
2674 CUUGGACA G CUCCUGGG 1592 CCCAGGAG GCCGAAAGGCGAGUGAGGUCU UGUCCAAG 3895
2704 AUGGAACA G CUCACAAG 1593 CUUGUGAG GCCGAAAGGCGAGUGAGGUCU UGUUCCAU 3896
2712 GCUCACAA G UAUAUCAU 1594 AUGAUAUA GCCGAAAGGCGAGUGAGGUCU UUGUGAGC 3897
2729 UCGAAUAA G UACAAGUA 1595 UACUUGUA GCCGAAAGGCGAGUGAGGUCU UUAUUCGA 3898
2735 AAGUACAA G UAUUCUUG 1596 CAAGAAUA GCCGAAAGGCGAGUGAGGUCU UUGUACUU 3899
2757 AGAGACAA G UUCAAUGA 1597 UCAUUGAA GCCGAAAGGCGAGUGAGGUCU UUGUCUCU 3900
2776 CUCUUCAA G UGAAUACU 1598 AGUAUUCA GCCGAAAGGCGAGUGAGGUCU UUGAAGAG 3901
2806 CAAAGGAA G CCAACUCU 1599 AGAGUUGG GCCGAAAGGCGAGUGAGGUCU UUCCUUUG 3902
2821 CUGAGGAA G UCUUUUUG 1600 CAAAAAGA GCCGAAAGGCGAGUGAGGUCU UUCCUCAG 3903
2861 UGAAAAUG G CACAGAUC 1601 GAUCUGUG GCCGAAAGGCGAGUGAGGUCU CAUUUUCA 3904
2887 CUAUUCAG G CUGUUGAU 1602 AUCAACAG GCCGAAAGGCGAGUGAGGUCU CUGAAUAG 3905
2899 UUGAUAAG G UCGAUCUG 1603 CAGAUCGA GCCGAAAGGCGAGUGAGGUCU CUUAUCAA 3906
2935 UUGCACGA G UAUCUUUG 1604 CAAAGAUA GCCGAAAGGCGAGUGAGGUCU UCGUGCAA 3907
2978 GACACCUA G UCCUGAUG 1605 CAUCAGGA GCCGAAAGGCGAGUGAGGUCU UAGGUGUC 3908
2991 GAUGAAAC G UCUGCUCC 1606 GGAGCAGA GCCGAAAGGCGAGUGAGGUCU GUUUCAUC 3909
3023 UAUCAACA G CACCAUUC 1607 GAAUGGUG GCCGAAAGGCGAGUGAGGUCU UGUUGAUA 3910
Figure imgf000102_0001
Input Sequence = NM_001285. Cut Site = G/Y
Arm Length = 8. Core Sequence = GCcgaaagGCGaGuCaaGGuCu
NM_001285 (Homo sapiens chloride channel, calcium activated, 1 (CLCAl) mRNA, 3311 bp)
Table VII: Human CLCAl DNAzyme and Target Sequence 249.021
Pos Substrate Seq ID DNAzyme Rz No Seq ID
No
17 CUUUUGGU A CAAAUGGA 4 TCCATTTG GGCTAGCTACAACGA ACCAAAAG 3919
34 UGUGGAAU A UAAUUGAA 5 TTCAATTA GGCTAGCTACAACGA ATTCCACA 3920
44 AAUUGAAU A UUUUCUUG 8 CAAGAAAA GGCTAGCTACAACGA ATTCAATT 3921
84 UUGAGGUU A UGUCAAGC 19 GCTTGACA GGCTAGCTACAACGA AACCTCAA 3922
122 AUGGAAAU A UUUACAAG 22 CTTGTAAA GGCTAGCTACAACGA ATTTCCAT 3923
126 AAAUAUUU A CAAGUACG 25 CGTACTTG GGCTAGCTACAACGA AAATATTT 3924
132 UUACAAGU A CGCAAUUU 26 AAATTGCG GGCTAGCTACAACGA ACTTGTAA 3925
152 ACUAAGAU A UUGUUAUC 30 GATAACAA GGCTAGCTACAACGA ATCTTAGT 3926
158 AUAUUGUU A UCAUUCUC 33 GAGAATGA GGCTAGCTACAACGA AACAATAT 3927
169 AUUCUCCU A UUGAAGAC 38 GTCTTCAA GGCTAGCTACAACGA AGGAGAAT 3928
259 GUGUGUCU A UAUUUUCA 52 TGAAAATA GGCTAGCTACAACGA AGACACAC 3929
261 GUGUCUAU A UUUUCAUA 53 TATGAAAA GGCTAGCTACAACGA ATAGACAC 3930
269 AUUUUCAU A UCUGUAUA 58 TATACAGA GGCTAGCTACAACGA ATGAAAAT 3931
275 AUAUCUGU A UAUAUAUA 60 TATATATA GGCTAGCTACAACGA ACAGATAT 3932
277 AUCUGUAU A UAUAUAAU 61 ATTATATA GGCTAGCTACAACGA ATACAGAT 3933
279 CUGUAUAU A UAUAAUGG 62 CCATTATA GGCTAGCTACAACGA ATATACAG 3934
281 GUAUAUAU A UAAUGGUA 63 TACCATTA GGCTAGCTACAACGA ATATATAC 3935
346 GGAGAUGU A CAGCAAUG 74 CATTGCTG GGCTAGCTACAACGA ACATCTCC 3936
446 CAAUGGCU A UGAAGGCA 97 TGCCTTCA GGCTAGCTACAACGA AGCCATTG 3937
539 AUCUCUGU A UCUGUUUG 108 CAAACAGA GGCTAGCTACAACGA ACAGAGATj 3938
553 UUGAAGCU A CAGGAAAG 112 CTTTCCTG GGCTAGCTACAACGA AGCTTCAA 3939
569 GCGAUUUU A UUUCAAAA 116 TTTTGAAA GGCTAGCTACAACGA AAAATCGC 3940
623 GGCUGACU A UGUGAGAC 126 GTCTCACA GGCTAGCTACAACGA AGTCAGCC 3941
647 UGAGACCU A CAAAAAUG 128 CATTTTTG GGCTAGCTACAACGA AGGTCTCA 3942
679 CUGAGUCU A CUCCUCCA 133 TGGAGGAG GGCTAGCTACAACGA AGACTCAG 3943
704 UGAACCCU A CACUGAGC 137 GCTCAGTG GGCTAGCTACAACGA AGGGTTCA 3944
791 AGCUGAAU A UGGACCAC 147 GTGGTCCA GGCTAGCTACAACGA ATTCAGCT 3945
834 GCUCAUCU A CGAUGGGG 154 CCCCATCG GGCTAGCTACAACGA AGATGAGC 3946
846 UGGGGAGU A UUUGACGA 155 TCGTCAAA GGCTAGCTACAACGA ACTCCCCA 3947
857 UGACGAGU A CAAUAAUG 158 CATTATTG GGCTAGCTACAACGA ACTCGTCA 3948
878 GAAAUUCU A CUUAUCCA 162 TGGATAAG GGCTAGCTACAACGA AGAATTTC 3949
882 UUCUACUU A UCCAAUGG 164 CCATTGGA GGCTAGCTACAACGA AAGTAGAA 3950
897 GGAAGAAU A CAAGCAGU 166 ACTGCTTG GGCTAGCTACAACGA ATTCTTCC 3951
922 CAGCAGGU A UUACUGGU 170 ACCAGTAA GGCTAGCTACAACGA ACCTGCTG 3952
925 CAGGUAUU A CUGGUACA 172 TGTACCAG GGCTAGCTACAACGA AATACCTG 3953
931 UUACUGGU A CAAAUGUA 173 TACATTTG GGCTAGCTACAACGA ACCAGTAA 3954
968 CAGCUGUU A CACCAAAA 178 TTTTGGTG GGCTAGCTACAACGA AACAGCTG 3955
997 AUAAAGUU A CAGGACUC 183 GAGTCCTG GGCTAGCTACAACGA AACTTTAT 3956
1007 AGGACUCU A UGAAAAAG 185 CTTTTTCA GGCTAGCTACAACGA AGAGTCCT 3957
1060 AGGCUUCU A UAAUGUUU 194 AAACATTA GGCTAGCTACAACGA AGAAGCCT 3958
1087 UUGAUUCU A UAGUUGAA 201 TTCAACTA GGCTAGCTACAACGA AGAATCAA 3959
1102 AAUUCUGU A CAGAACAA 206 TTGTTCTG GGCTAGCTACAACGA ACAGAATT 3960
1213 CCACUCCU A UGACAACA 218 TGTTGTCA GGCTAGCTACAACGA AGGAGTGG 3961
1416 GCCCAUGU A CAAAGUGA 245 TCACTTTG GGCTAGCTACAACGA ACATGGGC 3962
1431 GAACUCAU A CAGAUAAA 247 TTTATCTG GGCTAGCTACAACGA ATGAGTTC 3963
1476 AAAAGAUU A CCUGCAGC 251 GCTGCAGG GGCTAGCTACAACGA AATCTTTT 3964
1531 CGGCAUUU A CUGUGAUU 261 AATCACAG GGCTAGCTACAACGA AAATGCCG 3965
1550 GAAGAAAU A UCCAACUG 264 CAGTTGGA GGCTAGCTACAACGA ATTTCTTC 3966
1603 ACAACACU A UAAGUGGG 268 CCCACTTA GGCTAGCTACAACGA AGTGTTGT 3967
1716 GGAGGUUU A CAGACAUA 285 TATGTCTG GGCTAGCTACAACGA AAACCTCC 3968
1724 ACAGACAU A UGCUUCAG 286 CTGAAGCA GGCTAGCTACAACGA ATGTCTGT 3969 1909 UGUUUCUU A UCACCUGG 318 CCAGGTGA GGCTAGCTACAACGA AAGAAACA 3970
2006 AAUGGCCU A CCUCCAAA 329 TTTGGAGG GGCTAGCTACAACGA AGGCCATT 3971
2048 UUGGAAAU A CAGUCUGC 336 GCAGACTG GGCTAGCTACAACGA ATTTCCAA 3972
2110 CCAAUGCU A CCCUGCCU 343 AGGCAGGG GGCTAGCTACAACGA AGCATTGG 3973
2125 CUCCAAUU A CAGUGACU 346 AGTCACTG GGCTAGCTACAACGA AATTGGAG 3974
2183 GGUAGUUU A UGCAAAUA 355 TATTTGCA GGCTAGCTACAACGA AAACTACC 3975
2191 AUGCAAAU A UUCGCCAA 356 TTGGCGAA GGCTAGCTACAACGA ATTTGCAT 3976
2266 AAACAGUU A CCUUGGAA 367 TTCCAAGG GGCTAGCTACAACGA AACTGTTT 3977
2277 UUGGAACU A CUGGAUAA 369 TTATCCAG GGCTAGCTACAACGA AGTTCCAA 3978
2305 CUGAUGCU A CUAAGGAU 371 ATCCTTAG GGCTAGCTACAACGA AGCATCAG 3979
2324 CGGUGUCU A CUCAAGGU 374 ACCTTGAG GGCTAGCTACAACGA AGACACCG 3980
2333 CUCAAGGU A UUUCACAA 376 TTGTGAAA GGCTAGCTACAACGA ACCTTGAG 3981
2345 CACAACUU A UGACACGA 381 TCGTGTCA GGCTAGCTACAACGA AAGTTGTG 3982
2363 UGGUAGAU A CAGUGUAA 383 TTACACTG GGCTAGCTACAACGA ATCTACCA 3983
2418 AGAGUGAU A CCCCAGCA 388 TGCTGGGG GGCTAGCTACAACGA ATCACTCT 3984
2441 AGCACUGU A CAUACCUG 389 CAGGTATG GGCTAGCTACAACGA ACAGTGCT 3985
2445 CUGUACAU A CCUGGCUG 390 CAGCCAGG GGCTAGCTACAACGA ATGTACAG 3986
2472 GAUGAAAU A CAAUGGAA 392 TTCCATTG GGCTAGCTACAACGA ATTTCATC 3987
2592 GCUCCCAU A CCUGAUCU 411 AGATCAGG GGCTAGCTACAACGA ATGGGAGC 3988
2690 GGAUGAUU A UGACCAUG 427 CATGGTCA GGCTAGCTACAACGA AATCATCC 3989
2714 UCACAAGU A UAUCAUUC 429 GAATGATA GGCTAGCTACAACGA ACTTGTGA 3990
2716 ACAAGUAU A UCAUUCGA 430 TCGAATGA GGCTAGCTACAACGA ATACTTGT 3991
2731 GAAUAAGU A CAAGUAUU 435 AATACTTG GGCTAGCTACAACGA ACTTATTC 3992
2737 GUACAAGU A UUCUUGAU 436 ATCAAGAA GGCTAGCTACAACGA ACTTGTAC 3993
2782 AAGUGAAU A CUACUGCU 448 AGCAGTAG GGCTAGCTACAACGA ATTCACTT 3994
2785 UGAAUACU A CUGCUCUC 449 GAGAGCAG GGCTAGCTACAACGA AGTATTCA 3995
2848 AAAACAUU A CUUUUGAA 463 TTCAAAAG GGCTAGCTACAACGA AATGTTTT 3996
2881 UCAUUGCU A UUCAGGCU 473 AGCCTGAA GGCTAGCTACAACGA AGCAATGA 3997
2919 UCAGAAAU A UCCAACAU 481 ATGTTGGA GGCTAGCTACAACGA ATTTCTGA 3998
2937 GCACGAGU A UCUUUGUU 484 AACAAAGA GGCTAGCTACAACGA ACTCGTGC 3999
2947 CUUUGUUU A UUCCUCCA 490 TGGAGGAA GGCTAGCTACAACGA AAACAAAG 4000
3010 GUCCUAAU A UUCAUAUC 502 GATATGAA GGCTAGCTACAACGA ATTAGGAC 4001
3016 AUAUUCAU A UCAACAGC 505 GCTGTTGA GGCTAGCTACAACGA ATGAATAT 4002
3055 UAAAAAUU A UGUGGAAG 516 CTTCCACA GGCTAGCTACAACGA AATTTTTA 4003
3149 UUUUGAUU A UAAAAUUU 540 AAATTTTA GGCTAGCTACAACGA AATCAAAA 4004
3168 UAAAAUGU A UUUUAGAC 547 GTCTAAAA GGCTAGCTACAACGA ACATTTTA 4005
3194 GGGGCGAU A UACUAAAU 555 ATTTAGTA GGCTAGCTACAACGA ATCGCCCC 4006
3247 GGGGCGAU A UACUAAAU 555 ATTTAGTA GGCTAGCTACAACGA ATCGCCCC 4006
3196 GGCGAUAU A CUAAAUGU 556 ACATTTAG GGCTAGCTACAACGA ATATCGCC 4007
3249 GGCGAUAU A CUAAAUGU 556 ACATTTAG GGCTAGCTACAACGA ATATCGCC 4007
3205 CUAAAUGU A UAUAGUAC 558 GTACTATA GGCTAGCTACAACGA ACATTTAG 4008
3207 AAAUGUAU A UAGUACAU 559 ATGTACTA GGCTAGCTACAACGA ATACATTT 4009
3212 UAUAUAGU A CAUUUAUA 561 TATAAATG GGCTAGCTACAACGA ACTATATA 4010
3218 GUACAUUU A UACUAAAU 564 ATTTAGTA GGCTAGCTACAACGA AAATGTAC 4011
3220 ACAUUUAU A CUAAAUGU 565 ACATTTAG GGCTAGCTACAACGA ATAAATGT 4012
3229 CUAAAUGU A UUCCUGUA 567 TACAGGAA GGCTAGCTACAACGA ACATTTAG 4013
3258 CUAAAUGU A UUUUAGAC 572 GTCTAAAA GGCTAGCTACAACGA ACATTTAG 4014
65 AGGGGAGC A UGAAGAGG 579 CCTCTTCA GGCTAGCTACAACGA GCTCCCCT 4015
93 UGUCAAGC A UCUGGCAC 581 GTGCCAGA GGCTAGCTACAACGA GCTTGACA 4016
100 CAUCUGGC A CAGCUGAA 583 TTCAGCTG GGCTAGCTACAACGA GCCAGATG 4017
161 UUGUUAUC A UUCUCCUA 590 TAGGAGAA GGCTAGCTACAACGA GATAACAA 4018
195 AGUAAAAC A CAUCAGGU ■ 596 ACCTGATG GGCTAGCTACAACGA GTTTTACT 4019
197 UAAAACAC A UCAGGUCA 597 TGACCTGA GGCTAGCTACAACGA GTGTTTTA 4020
231 GAUAAACC A CUUCCGAU 603 ATCGGAAG GGCTAGCTACAACGA GGTTTATC 4021
267 AUAUUUUC A UAUCUGUA 607 TACAGATA GGCTAGCTACAACGA GAAAATAT 4022
299 AGAAAGAC A CCUUCGUA 609 • ACGAAGG GGCTAGCTACAACGA GTCTTTCT 4023 314 UAACCCGC A UUUUCCAA 614 TTGGAAAA GGCTAGCTACAACGA GCGGGTTA 4024
334 GAGGAAUC A CAGGGAGA 617 TCTCCCTG GGCTAGCTACAACGA GATTCCTC 4025
360 AUGGGGCC A UUUAAGAG 622 CTCTTAAA GGCTAGCTACAACGA GGCCCCAT 4026
379 CUGUGUUC A UCUUGAUU 624 AATCAAGA GGCTAGCTACAACGA GAACACAG 4027
392 GAUUCUUC A CCUUCUAG 627 CTAGAAGG GGCTAGCTACAACGA GAAGAATC 4028
420 AGUAAUUC A CUCAUUCA 634 TGAATGAG GGCTAGCTACAACGA GAATTACT 4029
424 AUUCACUC A UUCAGCUG 636 CAGCTGAA GGCTAGCTACAACGA GAGTGAAT 4030
454 AUGAAGGC A UUGUCGUU 642 AACGACAA GGCTAGCTACAACGA GCCTTCAT 4031
495 GAUGAAAC A CUCAUUCA 650 TGAATGAG GGCTAGCTACAACGA GTTTCATC 4032
499 AAACACUC A UUCAACAA 652 TTGTTGAA GGCTAGCTACAACGA GAGTGTTT 4033
517 UAAAGGAC A UGGUGACC 655 GGTCACCA GGCTAGCTACAACGA GTCCTTTA 4034
531 ACCCAGGC A UCUCUGUA 659 TACAGAGA GGCTAGCTACAACGA GCCTGGGT 4035
586 AUGUUGCC A UUUUGAUU 667 AATCAAAA GGCTAGCTACAACGA GGCAACAT 4036
603 CCUGAAAC A UGGAAGAC 670 GTCTTCCA GGCTAGCTACAACGA GTTTCAGG 4037
706 AACCCUAC A CUGAGCAG 692 CTGCTCAG GGCTAGCTACAACGA GTAGGGTT 4038
749 AAGGAUCC A CCUCACUC 698 GAGTGAGG GGCTAGCTACAACGA GGATCCTT 4039
754 UCCACCUC A CUCCUGAU 701 ATCAGGAG GGCTAGCTACAACGA GAGGTGGA 4040
766 CUGAUUUC A UUGCAGGA 705 TCCTGCAA GGCTAGCTACAACGA GAAATCAG 4041
798 UAUGGACC A CAAGGUAA 709 TTACCTTG GGCTAGCTACAACGA GGTCCATA 4042
810 GGUAAGGC A UUUGUCCA 711 TGGACAAA GGCTAGCTACAACGA GCCTTACC 4043
818 AUUUGUCC A UGAGUGGG 713 CCCACTCA GGCTAGCTACAACGA GGACAAAT 4044
830 GUGGGCUC A UCUACGAU 715 ATCGTAGA GGCTAGCTACAACGA GAGCCCAC 4045
970 GCUGUUAC A CCAAAAGA 731 TCTTTTGG GGCTAGCTACAACGA GTAACAGC 4046
982 AAAGAUGC A CAUUCAAU 734 ATTGAATG GGCTAGCTACAACGA GCATCTTT 4047
984 AGAUGCAC A UUCAAUAA 735 TTATTGAA GGCTAGCTACAACGA GTGCATCT 4048
1071 AUGUUUGC A CAACAUGU 749 ACATGTTG GGCTAGCTACAACGA GCAAACAT 4049
1076 UGCACAAC A UGUUGAUU 751 AATCAACA GGCTAGCTACAACGA GTTGTGCA 4050
1115 ACAAAACC A CAACAAAG 757 CTTTGTTG GGCTAGCTACAACGA GGTTTTGT 4051
1165 UCCGAAGC A CAUGGGAA 769 TTCCCATG GGCTAGCTACAACGA GCTTCGGA 4052
1167 CGAAGCAC A UGGGAAGU 770 ACTTCCCA GGCTAGCTACAACGA GTGCTTCG 4053
1207 AGAAAACC A CUCCUAUG 775 CATAGGAG GGCTAGCTACAACGA GGTTTTCT 4054
1221 AUGACAAC A CAGCCACC 780 GGTGGCTG GGCTAGCTACAACGA GTTGTCAT 4055
1227 ACACAGCC A CCAAAUCC 783 GGATTTGG GGCTAGCTACAACGA GGCTGTGT 4056
1237 CAAAUCCC A CCUUCUCA 788 TGAGAAGG GGCTAGCTACAACGA GGGATTTG 4057
1245 ACCUUCUC A UUGCUGCA 792 TGCAGCAA GGCTAGCTACAACGA GAGAAGGT 4058
1300 CUGGAAGC A UGGCGACU 800 AGTCGCCA GGCTAGCTACAACGA GCTTCCAG 4059
1395 AUGGUGAC A UUUGACAG 820 CTGTCAAA GGCTAGCTACAACGA GTCACCAT 4060
1412 UGCUGCCC A UGUACAAA 825 TTTGTACA GGCTAGCTACAACGA GGGCAGCA 4061
1429 GUGAACUC A UACAGAUA 828 TATCTGTA GGCTAGCTACAACGA GAGTTCAC 4062
1459 ACAGGGAC A CACUCGCC 833 GGCGAGTG GGCTAGCTACAACGA GTCCCTGT 4063
1461 AGGGACAC A CUCGCCAA 834 TTGGCGAG GGCTAGCTACAACGA GTGTCCCT 4064
1504 GGACGUCC A UCUGCAGC 845 GCTGCAGA GGCTAGCTACAACGA GGACGTCC 4065
1527 CGAUCGGC A UUUACUGU 849 ACAGTAAA GGCTAGCTACAACGA GCCGATCG 4066
1600 AAGACAAC A CUAUAAGU 858 ACTTATAG GGCTAGCTACAACGA GTTGTCTT 4067
1642 GUGGUGCC A UCAUCCAC 864 GTGGATGA GGCTAGCTACAACGA GGCACCAC 4068
1645 GUGCCAUC A UCCACACA 865 TGTGTGGA GGCTAGCTACAACGA GATGGCAC 4069
1649 CAUCAUCC A CACAGUCG 867 CGACTGTG GGCTAGCTACAACGA GGATGATG 4070
1651 UCAUCCAC A CAGUCGCU 868 AGCGACTG GGCTAGCTACAACGA GTGGATGA 4071
1722 UUACAGAC A UAUGCUUC 884 GAAGCATA GGCTAGCTACAACGA GTCTGTAA 4072
1756 AUGGCCUC A UUGAUGCU 892 AGCATCAA GGCTAGCTACAACGA GAGGCCAT 4073
1779 GCCCUUUC A UCAGGAAA 897 TTTCCTGA GGCTAGCTACAACGA GAAAGGGC 4074
1810 AGCGCUCC A UCCAGCUU 905 AAGCTGGA GGCTAGCTACAACGA GGAGCGCT 4075
1864 UGAAUGGC A CAGUGAUC 917 GATCACTG GGCTAGCTACAACGA GCCATTCA 4076
1882 UGGACAGC A CCGUGGGA 920 TCCCACGG GGCTAGCTACAACGA GCTGTCCA 4077
1897 GAAAGGAC A CUUUGUUU 922 AAACAAAG GGCTAGCTACAACGA GTCCTTTC 4078
1912 UUCUUAUC A CCUGGACA 925 TGTCCAGG GGCTAGCTACAACGA GATAAGAA 4079 1993 ACAAAAAC A CCAAAAUG 947 CATTTTGG GGCTAGCTACAACGA GTTTTTGT 4080
2023 UCCCAGGC A UUGCUAAG 959 CTTAGCAA GGCTAGCTACAACGA GCCTGGGA 4081
2038 AGGUUGGC A CUUGGAAA 961 TTTCCAAG GGCTAGCTACAACGA GCCAACCT 4082
2067 GCAAGCUC A CAAACCUU 968 AAGGTTTG GGCTAGCTACAACGA GAGCTTGC 4083
2089 UGACUGUC A CGUCCCGU 976 ACGGGACG GGCTAGCTACAACGA GACAGTCA 4084
2152 ACAAGGAC A CCAGCAAA 994 TTTGCTGG GGCTAGCTACAACGA GTCCTTGT 4085
2230 CCAGUGUC A CAGCCCUG 1019 CAGGGCTG GGCTAGCTACAACGA GACACTGG 4086
2338 GGUAUUUC A CAACUUAU 1037 ATAAGTTG GGCTAGCTACAACGA GAAATACC 4087
2350 CUUAUGAC A CGAAUGGU 1040 ACCATTCG GGCTAGCTACAACGA GTCATAAG 4088
2436 AGUGGAGC A CUGUACAU 1052 ATGTACAG GGCTAGCTACAACGA GCTCCACT 4089
2443 CACUGUAC A UACCUGGC 1054 GCCAGGTA GGCTAGCTACAACGA GTACAGTG 4090
2484 UGGAAUCC A CCAAGACC 1060 GGTCTTGG GGCTAGCTACAACGA GGATTCCA 4091
2519 UGUUCAAC A CAAGCAAG 1066 CTTGCTTG GGCTAGCTACAACGA GTTGAACA 4092
2544 AGCAGAAC A UCCUCGGG 1071 CCCGAGGA GGCTAGCTACAACGA GTTCTGCT 4093
2559 GGAGGCUC A UUUGUGGC 1075 GCCACAAA GGCTAGCTACAACGA GAGCCTCC 4094
2590 AUGCUCCC A UACCUGAU 1084 ATCAGGTA GGCTAGCTACAACGA GGGAGCAT 4095
2607 CUCUUCCC A CCUGGCCA 1091 TGGCCAGG GGCTAGCTACAACGA GGGAAGAG 4096
2620 GCCAAAUC A CCGACCUG 1096 CAGGTCGG GGCTAGCTACAACGA GATTTGGC 4097
2642 GGAAAUUC A CGGGGGCA 1100 TGCCCCCG GGCTAGCTACAACGA GAATTTCC 4098
2656 GCAGUCUC A UUAAUCUG 1103 CAGATTAA GGCTAGCTACAACGA GAGACTGC 4099
2696 UUAUGACC A UGGAACAG 1111 CTGTTCCA GGCTAGCTACAACGA GGTCATAA 4100
2708 AACAGCUG A CAAGUAUA 1114 TATACTTG GGCTAGCTACAACGA GAGCTGTT 4101
2719 AGUAUAUC A UUCGAAUA 1116 TATTCGAA GGCTAGCTACAACGA GATATACT 4102
2794 CUGCUCUC A UCCCAAAG 1130 CTTTGGGA GGCTAGCTACAACGA GAGAGCAG 4103
2845 CAGAAAAC A UUACUUUU 1141 AAAAGTAA GGCTAGCTACAACGA GTTTTCTG 4104
2863 AAAAUGGC A CAGAUCUU 1143 AAGATCTG GGCTAGCTACAACGA GCCATTTT 4105
2875 AUCUUUUC A UUGCUAUU 1146 AATAGCAA GGCTAGCTACAACGA GAAAAGAT 4106
2926 UAUCCAAC A UUGCACGA 1154 TCGTGCAA GGCTAGCTACAACGA GTTGGATA 4107
2931 AACAUUGC A CGAGUAUC 1155 GATACTCG GGCTAGCTACAACGA GCAATGTT 4108
2955 AUUCCUCC A CAGACUCC 1160 GGAGTCTG GGCTAGCTACAACGA GGAGGAAT 4109
2973 CCAGAGAC A CCUAGUCC 1166 GGACTAGG GGCTAGCTACAACGA GTCTCTGG 4110
3014 UAAUAUUC A UAUCAACA 1177 TGTTGATA GGCTAGCTACAACGA GAATATTA 4111
3025 UCAACAGC A CCAUUCCU 1180 AGGAATGG GGCTAGCTACAACGA GCTGTTGA 4112
3028 ACAGCACC A UUCCUGGC 1182 GCCAGGAA GGCTAGCTACAACGA GGTGCTGT 4113
3037 UUCCUGGC A UUCACAUU 1185 AATGTGAA GGCTAGCTACAACGA GCCAGGAA 4114
3041 UGGCAUUC A CAUUUUAA 1186 TTAAAATG GGCTAGCTACAACGA GAATGCCA 4115
3043 GCAUUCAC A UUUUAAAA 1187 TTTTAAAA GGCTAGCTACAACGA GTGAATGC 4116
3130 AAUAAAUC A UUCAUCCU 1196 AGGATGAA GGCTAGCTACAACGA GATTTATT 4117
3134 AAUCAUUC A UCCUUUUU 1197 AAAAAGGA GGCTAGCTACAACGA GAATGATT 4118
3214 UAUAGUAC A UUUAUACU 1205 AGTATAAA GGCTAGCTACAACGA GTACTATA 4119
134 ACAAGUAC G CAAUUUGA 1215 TCAAATTG GGCTAGCTACAACGA .GTACTTGT 4120
312 CGUAACCC G CAUUUUCC 1220 GGAAAATG GGCTAGCTACAACGA GGGTTACG 4121
463 UUGUCGUU G CAAUCGAC 1225 GTCGATTG GGCTAGCTACAACGA AACGACAA 4122
480 CCCAAUGU G CCAGAAGA 1227 TCTTCTGG GGCTAGCTACAACGA ACATTGGG 4123
583 AAAAUGUU G CCAUUUUG 1232 CAAAATGG GGCTAGCTACAACGA AACATTTT 4124
655 ACAAAAAU G CUGAUGUU 1238 AACATCAG GGCTAGCTACAACGA ATTTTTGT 4125
670 UUCUGGUU G CUGAGUCU 1240 AGACTCAG GGCTAGCTACAACGA AACCAGAA 4126
769 AUUUCAUU G CAGGAAAA 1247 TTTTCCTG GGCTAGCTACAACGA AATGAAAT 4127
980 CAAAAGAU G CACAUUCA 1255 TGAATGTG GGCTAGCTACAACGA ATCTTTTG 4128
1040 CCAAUCCC G CCAGACGG 1258 CCGTCTGG GGCTAGCTACAACGA GGGATTGG 4129
1069 UAAUGUUU G CACAACAU 1259 ATGTTGTG GGCTAGCTACAACGA AAACATTA 4130
1151 UCAAAAAU G CAAUCUCC 1262 GGAGATTG GGCTAGCTACAACGA ATTTTTGA 4131
1248 UUCUCAUU G CUGCAGAU 1268 ATCTGCAG GGCTAGCTACAACGA AATGAGAA 4132
1251 UCAUUGCU G CAGAUUGG 1269 CCAATCTG GGCTAGCTACAACGA AGCAATGA 4133
1316 UGGUAACC G CCUCAAUC 1272 GATTGAGG GGCTAGCTACAACGA GGTTACCA 4134
1353 CUUUUCCU G CUGCAGAC 1275 GTCTGCAG GGCTAGCTACAACGA AGGAAAAG 4135 1356 UUCCUGCU I cAGACAGU 1276 ACTGTCTG GGCTAGCTACAACGA AGCAGGAA 4136
1405 UUGACAGU G CUGCCCAU 1280 ATGGGCAG GGCTAGCTACAACGA ACTGTCAA 4137
1408 ACAGUGCU G CCCAUGUA 1281 TACATGGG GGCTAGCTACAACGA AGCACTGT 4138
1465 ACACACUC G CCAAAAGA 1284 TCTTTTGG GGCTAGCTACAACGA GAGTGTGT 4139
1480 GAUUACCU G CAGCAGCU 1285 AGCTGCTG GGCTAGCTACAACGA AGGTAATC 4140
1508 GUCCAUCU G CAGCGGGC 1286 GCCCGCTG GGCTAGCTACAACGA AGATGGAC 4141
1575 GAAAUUGU G CUGCUGAC 1291 GTCAGCAG GGCTAGCTACAACGA ACAATTTC 4142
1578 AUUGUGCU G CUGACGGA 1292 TCCGTCAG GGCTAGCTACAACGA AGCACAAT 4143
1613 AAGUGGGU G CUUUAACG 1294 CGTTAAAG GGCTAGCTACAACGA ACCCACTT 4144
1639 AAAGUGGU G CCAUCAUC 1296 GATGATGG GGCTAGCTACAACGA ACCACTTT 4145
1657 ACACAGUC G CUUUGGGG 1297 CCCCAAAG GGCTAGCTACAACGA GACTGTGT 4146
1672 GGCCCUCU G CAGCUCAA 1298 TTGAGCTG GGCTAGCTACAACGA AGAGGGCC 4147
1726 AGACAUAU G CUUCAGAU 1300 ATCTGAAG GGCTAGCTACAACGA ATATGTCT 4148
1762 UCAUUGAU G CUUUUGGG 1302 CCCAAAAG GGCTAGCTACAACGA ATCAATGA 4149
1805 CUCUCAGC G CUCCAUCC 1303 GGATGGAG GGCTAGCTACAACGA GCTGAGAG 4150
1923 UGGACAAC G CAGCCUCC 1307 GGAGGCTG GGCTAGCTACAACGA GTTGTCCA 4151
2026 CAGGCAUU G CUAAGGUU 1308 AACCTTAG GGCTAGCTACAACGA AATGCCTG 4152
2055 UACAGUCU G CAAGCAAG 1309 CTTGCTTG GGCTAGCTACAACGA AGACTGTA 4153
2098 CGUCCCGU G CGUCCAAU 1312 ATTGGACG GGCTAGCTACAACGA ACGGGACG 4154
2107 CGUCCAAU G CUACCCUG 1313 CAGGGTAG GGCTAGCTACAACGA ATTGGACG 4155
2115 GCUACCCU G CCUCCAAU 1314 ATTGGAGG GGCTAGCTACAACGA AGGGTAGC 4156
2185 UAGUUUAU G CAAAUAUU 1317 AATATTTG GGCTAGCTACAACGA ATAAACTA 4157
2195 AAAUAUUC G CCAAGGAG 1318 CTCCTTGG GGCTAGCTACAACGA GAATATTT 4158
2296 GAGCAGGU G CUGAUGCU 1322 AGCATCAG GGCTAGCTACAACGA ACCTGCTC 4159
2302 GUGCUGAU G CUACUAAG 1324 CTTAGTAG GGCTAGCTACAACGA ATCAGCAC 4160
2376_j GUAAAAGU G CGGGCUCU 1328 AGAGCCCG GGCTAGCTACAACGA ACTTTTAC 4161
2398 GAGUUAAC G CAGCCAGA 1329 TCTGGCTG GGCTAGCTACAACGA GTTAACTC 4162
2584 UCCCAAAU G CUCCCAUA 1337 TATGGGAG GGCTAGCTACAACGA ATTTGGGA 4163
2788 AUACUACU G CUCUCAUC 1348 GATGAGAG GGCTAGCTACAACGA AGTAGTAT 4164
2878 UUUUCAUU G CUAUUCAG 1351 CTGAATAG GGCTAGCTACAACGA AATGAAAA 4165
2929 CCAACAUU G CACGAGUA 1355 TACTCGTG GGCTAGCTACAACGA AATGTTGG 4166
2964 CAGACUCC G CCAGAGAC 1357 GTCTCTGG GGCTAGCTACAACGA GGAGTCTG 4167
2995 AAACGUCU G CUCCUUGU 1360 ACAAGGAG GGCTAGCTACAACGA AGACGTTT 4168
3078^ GGAGAACU G GAGCUGUC 1361 GACAGCTG GGCTAGCTACAACGA AGTTCTCC 4169
3294 AAUAAAAU G CUAAACAA 1366 TTGTTTAG GGCTAGCTACAACGA ATTTTATT 4170
27 AAAUGGAU G UGGAAUAU 1367 ATATTCCA GGCTAGCTACAACGA ATCCATTT 4171
52 AUUUUCUU G UUUAAGGG 1368 CCCTTAAA GGCTAGCTACAACGA AAGAAAAT 4172
75 GAAGAGGU G UUGAGGUU 1369 AACCTCAA GGCTAGCTACAACGA ACCTCTTC 4173
86 GAGGUUAU G UCAAGCAU 1370 ATGCTTGA GGCTAGCTACAACGA ATAACCTC 4174
155 AAGAUAUU G UUAUCAUU 1371 AATGATAA GGCTAGCTACAACGA AATATCTT 4175
221 AAAGACCU G UGAUAAAC 1372 GTTTATCA GGCTAGCTACAACGA AGGTCTTT 4176
253 GGAAACGU G UGUCUAUA 1373 TATAGACA GGCTAGCTACAACGA ACGTTTCC 4177
255 AAACGUGU G UCUAUAUU 1374 AATATAGA GGCTAGCTACAACGA ACACGTTT 4178
273 UCAUAUCU G UAUAUAUA 1375 TATATATA GGCTAGCTACAACGA AGATATGA 4179
344 AGGGAGAU G UACAGCAA 1376 TTGCTGTA GGCTAGCTACAACGA ATCTCCCT 4180
373 AGAGUUCU G UGUUCAUC 1377 GATGAACA GGCTAGCTACAACGA AGAACTCT 4181
375 AGUUCUGU G UUCAUCUU 1378 AAGATGAA GGCTAGCTACAACGA ACAGAACT 4182
457 AAGGCAUU G UCGUUGCA 1379 TGCAACGA GGCTAGCTACAACGA AATGCCTT 4183,
478 ACCCCAAU G UGCCAGAA 1380 TTCTGGCA GGCTAGCTACAACGA ATTGGGGT 4184
537 GCAUCUCU G UAUCUGUU 1381 AACAGATA GGCTAGCTACAACGA AGAGATGC 4185
543 CUGUAUCU G UUUGAAGC 1382 GCTTCAAA GGCTAGCTACAACGA AGATACAG 4186
580 UCAAAAAU G UUGCCAUU 1383 AATGGCAA GGCTAGCTACAACGA ATTTTTGA 4187
625 CUGACUAU G UGAGACCA 1384 TGGTCTCA GGCTAGCTACAACGA ATAGTCAG 4188
661 AUGCUGAU G UUCUGGUU 1385 AACCAGAA GGCTAGCTACAACGA ATCAGCAT 4189
725 GGGCAACU G UGGAGAGA 1386 TCTCTCCA GGCTAGCTACAACGA AGTTGCCC 4190
814 AGGCAUUU G UCCAUGAG 1387 CTCATGGA GGCTAGCTACAACGA AAATGCCT 4191 911 AGUAAGAU G UUCAGCAG 1388 CTGCTGAA GGCTAGCTACAACGA ATCTTACT 4192
937 GUACAAAU G UAGUAAAG 1389 CTTTACTA GGCTAGCTACAACGA ATTTGTAC 4193
950 AAAGAAGU G UCAGGGAG 1390 CTCCCTGA GGCTAGCTACAACGA ACTTCTTT 4194
965 AGGCAGCU G UUACACCA 1391 TGGTGTAA GGCTAGCTACAACGA AGCTGCCT 4195
1019 AAAAGGAU G UGAGUUUG 1392 CAAACTCA GGCTAGCTACAACGA ATCCTTTT 4196
1027 GUGAGUUU G UUCUCCAA 1393 TTGGAGAA GGCTAGCTACAACGA AAACTCAC 4197
1065 UCUAUAAU G UUUGCACA 1394 TGTGCAAA GGCTAGCTACAACGA ATTATAGA 4198
1078 CACAACAU G UUGAUUCU 1395 AGAATCAA GGCTAGCTACAACGA ATGTTGTG 4199
1100 UGAAUUCU G UACAGAAC 1396 GTTCTGTA GGCTAGCTACAACGA AGAATTCA 4200
1270 AAAGAAUU G UGUGUUUA 1397 TAAACACA GGCTAGCTACAACGA AATTCTTT 4201
1272 AGAAUUGU G UGUUUAGU 1398 ACTAAACA GGCTAGCTACAACGA ACAATTCT 4202
1274 AAUUGUGU G UUUAGUCC 1399 GGACTAAA GGCTAGCTACAACGA ACACAATT 4203
1414 CUGCCCAU G UACAAAGU 1400 ACTTTGTA GGCTAGCTACAACGA ATGGGCAG 4204
1534 CAUUUACU G UGAUUAGG 1401 CCTAATCA GGCTAGCTACAACGA AGTAAATG 4205
1573 CUGAAAUU G UGCUGCUG 1402 CAGCAGCA GGCTAGCTACAACGA AATTTCAG 4206
1695 GAGGAGCU G UCCAAAAU 1403 ATTTTGGA GGCTAGCTACAACGA AGCTCCTC 4207
1795 AUGGAGCU G UCUCUCAG 1404 CTGAGAGA GGCTAGCTACAACGA AGCTCCAT 4208
1902 GACACUUU G UUUCUUAU 1405 ATAAGAAA GGCTAGCTACAACGA AAAGTGTC 4209
1978 GUGGCUUU G UAGUGGAC 1406 GTCCACTA GGCTAGCTACAACGA AAAGCCAC 4210
2086 CCCUGACU G UCACGUCC 1407 GGACGTGA GGCTAGCTACAACGA AGTCAGGG 4211
2227 GGGCCAGU G UCACAGCC 1408 GGCTGTGA GGCTAGCTACAACGA ACTGGCCC 4212
2320 AUGACGGU G UCUACUCA 1409 TGAGTAGA GGCTAGCTACAACGA ACCGTCAT 4213
2368 GAUACAGU G UAAAAGUG 1410 CACTTTTA GGCTAGCTACAACGA ACTGTATC 4214
2439 GGAGCACU G UACAUACC 1411 GGTATGTA GGCTAGCTACAACGA AGTGCTCC 4215
2512 AGGAUGAU G UUCAACAC 1412 GTGTTGAA GGCTAGCTACAACGA ATCATCCT 4216
2529 AAGCAAGU G UGUUUCAG 1413 CTGAAACA GGCTAGCTACAACGA ACTTGCTT 4217
2531 GCAAGUGU G UUUCAGCA 1414 TGCTGAAA GGCTAGCTACAACGA ACACTTGC 4218
2563 GCUCAUUU G UGGCUUCU 1415 AGAAGCCA GGCTAGCTACAACGA AAATGAGC 4219
2575 CUUCUGAU G UCCCAAAU 1416 ATTTGGGA GGCTAGCTACAACGA ATCAGAAG 4220
2829 GUCUUUUU G UUUAAACC 1417 GGTTTAAA GGCTAGCTACAACGA AAAAAGAC 4221
2890 UUCAGGCU G UUGAUAAG 1418 CTTATCAA GGCTAGCTACAACGA AGCCTGAA 4222
2943 GUAUCUUU G UUUAUUCC 1419 GGAATAAA GGCTAGCTACAACGA AAAGATAC 4223
3002 UGCUCCUU G UCCUAAUA 1420 TATTAGGA GGCTAGCTACAACGA AAGGAGCA 4224
3057 AAAAUUAU G UGGAAGUG 1421 CACTTCCA GGCTAGCTACAACGA ATAATTTT 4225
3084 CUGCAGCU G UCAAUAGC 1422 GCTATTGA GGCTAGCTACAACGA AGCTGCAG 4226
3109 GAAUUUUU G UCAGAUAA 1423 TTATCTGA GGCTAGCTACAACGA AAAAATTC 4227
3166 UCUAAAAU G UAUUUUAG 1424 CTAAAATA GGCTAGCTACAACGA ATTTTAGA 4228
3182 GACUUCCU G UAGGGGGC 1425 GCCCCCTA GGCTAGCTACAACGA AGGAAGTC 4229
3272 GACUUCCU G UAGGGGGC 1425 GCCCCCTA GGCTAGCTACAACGA AGGAAGTC 4229
3203 UACUAAAU G UAUAUAGU 1426 ACTATATA GGCTAGCTACAACGA ATTTAGTA 4230
3227 UACUAAAU G UAUUCCUG 1427 CAGGAATA GGCTAGCTACAACGA ATTTAGTA 4231
3235 GUAUUCCU G UAGGGGGC 1428 GCCCCCTA GGCTAGCTACAACGA AGGAATAC 4232
3256 UACUAAAU G UAUUUUAG 1429 CTAAAATA GGCTAGCTACAACGA ATTTAGTA 4233
15 UGCUUUUG G UACAAAUG 1430 CATTTGTA GGCTAGCTACAACGA CAAAAGCA 4234
63 UAAGGGGA G CAUGAAGA 1431 TCTTCATG GGCTAGCTACAACGA TCCCCTTA 4235
73 AUGAAGAG G UGUUGAGG 1432 CCTCAACA GGCTAGCTACAACGA CTCTTCAT 4236
81 GUGUUGAG G UUAUGUCA 1433 TGACATAA GGCTAGCTACAACGA CTCAACAC 4237
91 UAUGUCAA G CAUCUGGC 1434 GCCAGATG GGCTAGCTACAACGA TTGACATA 4238
98 AGCAUCUG G CACAGCUG 1435 CAGCTGTG GGCTAGCTACAACGA CAGATGCT 4239
103 CUGGCACA G CUGAAGGC 1436 GCCTTCAG GGCTAGCTACAACGA TGTGCCAG 4240
110 AGCUGAAG G CAGAUGGA 1437 TCCATCTG GGCTAGCTACAACGA CTTCAGCT 4241
130 AUUUACAA G UACGCAAU 1438 ATTGCGTA GGCTAGCTACAACGA TTGTAAAT 4242
182 AGACAAGA G CAAUAGUA 1439 TACTATTG GGCTAGCTACAACGA TCTTGTCT 4243
188 GAGCAAUA G UAAAACAC 1440 GTGTTTTA GGCTAGCTACAACGA TATTGCTC 4244
202 CACAUCAG G UCAGGGGG 1441 CCCCCTGA GGCTAGCTACAACGA CTGATGTG 4245
210 GUCAGGGG G UUAAAGAC 1442 GTCTTTAA GGCTAGCTACAACGA CCCCTGAC 4246 242 UCCGAUAA G UUGGAAAC 1443 GTTTCCAA GGCTAGCTACAACGA TTATCGGA 4247
251 UUGGAAAC G UGUGUCUA 1444 TAGACACA GGCTAGCTACAACGA GTTTCCAA 4248
287 AUAUAAUG G UAAAGAAA 1445 TTTCTTTA GGCTAGCTACAACGA CATTATAT 4249
305 ACACCUUC G UAACCCGC 1446 GCGGGTTA GGCTAGCTACAACGA GAAGGTGT 4250
349 GAUGUACA G CAAUGGGG 1447 CCCCATTG GGCTAGCTACAACGA TGTACATC 4251
357 GCAAUGGG G CCAUUUAA 1448 TTAAATGG GGCTAGCTACAACGA CCCATTGC 4252
368 AUUUAAGA G UUCUGUGU 1449 ACACAGAA GGCTAGCTACAACGA TCTTAAAT 4253
406 UAGAAGGG G CCCUGAGU 1450 ACTCAGGG GGCTAGCTACAACGA CCCTTCTA 4254
413 GGCCCUGA G UAAUUCAC 1451 GTGAATTA GGCTAGCTACAACGA TCAGGGCC 4255
429 CUCAUUCA G CUGAACAA 1452 TTGTTCAG GGCTAGCTACAACGA TGAATGAG 4256
443 CAACAAUG G CUAUGAAG 1453 CTTCATAG GGCTAGCTACAACGA CATTGTTG 4257
452 CUAUGAAG G CAUUGUCG 1454 CGACAATG GGCTAGCTACAACGA CTTCATAG 4258
460 GCAUUGUC G UUGCAAUC 1455 GATTGCAA GGCTAGCTACAACGA GACAATGC 4259
520 AGGACAUG G UGACCCAG 1456 CTGGGTCA GGCTAGCTACAACGA CATGTCCT 4260
529 UGACCCAG G CAUCUCUG 1457 CAGAGATG GGCTAGCTACAACGA CTGGGTCA 4261
550 UGUUUGAA G CUACAGGA 1458 TCCTGTAG GGCTAGCTACAACGA TTCAAACA 4262
561 ACAGGAAA G CGAUUUUA 1459 TAAAATCG GGCTAGCTACAACGA TTTCCTGT 4263
616 AGACAAAG G CUGACUAU 1460 ATAGTCAG GGCTAGCTACAACGA CTTTGTCT 4264
667 AUGUUGUG G UUGCUGAG 1461 CTCAGCAA GGCTAGCTACAACGA CAGAACAT 4265
675 GUUGCUGA G UCUACUCC 1462 GGAGTAGA GGCTAGCTACAACGA TCAGCAAC 4266
689 UCCUCCAG G UAAUGAUG 1463 CATCATTA GGCTAGCTACAACGA CTGGAGGA 4267
711 UACACUGA G CAGAUGGG 1464 CCCATCTG GGCTAGCTACAACGA TCAGTGTA 4268
719 GCAGAUGG G CAACUGUG 1465 CACAGTTG GGCTAGCTACAACGA CCATCTGC 4269
737 AGAGAAGG G UGAAAGGA 1466 TCCTTTCA GGCTAGCTACAACGA CCTTCTCT 4270
780 GGAAAAAA G UUAGCUGA 1467 TCAGCTAA GGCTAGCTACAACGA TTTTTTCC 4271
784 AAAAGUUA G CUGAAUAU 1468 ATATTCAG GGCTAGCTACAACGA TAACTTTT 4272
803 ACCACAAG G UAAGGCAU •1469 ATGCCTTA GGCTAGCTACAACGA CTTGTGGT 4273
808 AAGGUAAG G CAUUUGUC 1470 GACAAATG GGCTAGCTACAACGA CTTACCTT 4274
822 GUCCAUGA G UGGGCUCA 1471 TGAGCCCA GGCTAGCTACAACGA TCATGGAC 4275
826 AUGAGUGG G CUCAUCUA 1472 TAGATGAG GGCTAGCTACAACGA CCACTCAT 4276
844_^ GAUGGGGA G UAUUUGAC 1473 GTCAAATA GGCTAGCTACAACGA TCCCCATC 4277
855 UUUGACGA G UACAAUAA 1474 TTATTGTA GGCTAGCTACAACGA TCGTCAAA 4278
901 GAAUACAA G CAGUAAGA 1475 TCTTACTG GGCTAGCTACAACGA TTGTATTC 4279
904 UACAAGCA G UAAGAUGU 1476 ACATCTTA GGCTAGCTACAACGA TGCTTGTA 4280
916 GAUGUUCA G CAGGUAUU 1477 AATACCTG GGCTAGCTACAACGA TGAACATC 4281
920 UUCAGCAG G UAUUACUG 1478 CAGTAATA GGCTAGCTACAACGA CTGCTGAA 4282
929 UAUUACUG G UACAAAUG 1479 CATTTGTA GGCTAGCTACAACGA CAGTAATA 4283
940 CAAAUGUA G UAAAGAAG 1480 CTTCTTTA GGCTAGCTACAACGA TACATTTG 4284
948 GUAAAGAA G UGUCAGGG 1481 CCCTGACA GGCTAGCTACAACGA TTCTTTAC 4285
959 UCAGGGAG G CAGCUGUU 1482 AACAGCTG GGCTAGCTACAACGA CTCCCTGA 4286
962 GGGAGGCA G CUGUUACA 1483 TGTAACAG GGCTAGCTACAACGA TGCCTCCC 4287
994 UCAAUAAA G UUACAGGA 1484 TCCTGTAA GGCTAGCTACAACGA TTTATTGA 4288
1023 GGAUGUGA G UUUGUUCU 1485 AGAACAAA GGCTAGCTACAACGA TCACATCC 4289
1054 CGGAGAAG G CUUCUAUA 1486 TATAGAAG GGCTAGCTACAACGA CTTCTCCG 4290
1090 AUUCUAUA G UUGAAUUC 1487 GAATTCAA GGCTAGCTACAACGA TATAGAAT 4291
1126 ACAAAGAA G CUCCAAAC 1488 GTTTGGAG GGCTAGCTACAACGA TTCTTTGT 4292
1137 CCAAACAA G CAAAAUCA 1489 TGATTTTG GGCTAGCTACAACGA TTGTTTGG 4293
1163 UCUCCGAA G CACAUGGG 1490 CCCATGTG GGCTAGCTACAACGA TTCGGAGA 4294
1174 CAUGGGAA G UGAUCCGU 1491 ACGGATCA GGCTAGCTACAACGA TTCCCATG 4295
1181 AGUGAUCC G UGAUUCUG 1492 CAGAATCA GGCTAGCTACAACGA GGATCACT 4296
1224 ACAACACA G CCACCAAA 1493 TTTGGTGG GGCTAGCTACAACGA TGTGTTGT 4297
1279 UGUGUUUA G UCCUUGAC 1494 GTCAAGGA GGCTAGCTACAACGA TAAACACA 4298
1298 AUCUGGAA G CAUGGCGA 1495 TCGCCATG GGCTAGCTACAACGA TTCCAGAT 4299
1303 GAAGCAUG G CGACUGGU 1496 ACCAGTCG GGCTAGCTACAACGA CATGCTTC 4300
1310 GGCGACUG G UAACCGCC 1497 GGCGGTTA GGCTAGCTACAACGA CAGTCGCC 4301
1336 UGAAUCAA G CAGGCCAG 1498 CTGGCCTG GGCTAGCTACAACGA TTGATTCA 4302 1340 UCAAGCAG G CCAGCUUU 1499 AAAGCTGG GGCTAGCTACAACGA CTGCTTGA 4303
1344 GCAGGCCA G CUUUUCCU 1500 AGGAAAAG GGCTAGCTACAACGA TGGCCTGC 4304
1363 UGCAGACA G UUGAGCUG 1501 CAGCTCAA GGCTAGCTACAACGA TGTCTGCA 4305
1368 ACAGUUGA G CUGGGGUC 1502 GACCCCAG GGCTAGCTACAACGA >TCAACTGT 4306
1374 GAGCUGGG G UCCUGGGU 1503 ACCCAGGA GGCTAGCTACAACGA CCCAGCTC 4307
1381 GGUCCUGG G UUGGGAUG 1504 CATCCCAA GGCTAGCTACAACGA CCAGGACC 4308
1390 UUGGGAUG G UGACAUUU 1505 AAATGTCA GGCTAGCTACAACGA CATCCCAA 4309
1403 AUUUGACA G UGCUGCCC 1506 GGGCAGCA GGCTAGCTACAACGA TGTCAAAT 4310
1421 UGUACAAA G UGAACUCA 1507 TGAGTTCA GGCTAGCTACAACGA TTTGTACA 4311
1442 GAUAAACA G UGGCAGUG 1508 CACTGCCA GGCTAGCTACAACGA TGTTTATC 4312
1445 AAACAGUG G CAGUGACA 1509 TGTCACTG GGCTAGCTACAACGA CACTGTTT 4313
1448 CAGUGGCA G UGACAGGG 1510 CCCTGTCA GGCTAGCTACAACGA TGCCACTG 4314
1483 UACCUGCA G CAGCUUCA 1511 TGAAGCTG GGCTAGCTACAACGA TGCAGGTA 4315
1486 CUGCAGCA G CUUCAGGA 1512 TCCTGAAG GGCTAGCTACAACGA TGCTGCAG 4316
1500 GGAGGGAC G UCCAUCUG 1513 CAGATGGA GGCTAGCTACAACGA GTCCCTCC 4317
1511 CAUCUGCA G CGGGCUUC 1514 GAAGCCCG GGCTAGCTACAACGA TGCAGATG 4318
1515 UGCAGCGG G CUUCGAUC 1515 GATCGAAG GGCTAGCTACAACGA CCGCTGCA 4319
1525 UUCGAUCG G CAUUUACU 1516 AGTAAATG GGCTAGCTACAACGA CGATCGAA 4320
1607 CACUAUAA G UGGGUGCU 1517 AGCACCCA GGCTAGCTACAACGA TTATAGTG 4321
1611 AUAAGUGG G UGCUUUAA 1518 TTAAAGCA GGCTAGCTACAACGA CCACTTAT 4322
1624 UUAACGAG G UCAAACAA 1519] TTGTTTGA GGCTAGCTACAACGA CTCGTTAA 4323
1634 CAAACAAA G UGGUGCCA 1520 TGGCACCA GGCTAGCTACAACGA TTTGTTTG 4324
1637 ACAAAGUG G UGCCAUCA 1521 TGATGGCA GGCTAGCTACAACGA CACTTTGT 4325
1654 UCCACACA G UCGCUUUG 1522 CAAAGCGA GGCTAGCTACAACGA TGTGTGGA 4326
1665 GCUUUGGG G CCCUCUGC 1523 GCAGAGGG GGCTAGCTACAACGA CCCAAAGC 4327
1675 CCUCUGCA G CUCAAGAA 1524 TTCTTGAG GGCTAGCTACAACGA TGCAGAGG 4328
1692 CUAGAGGA G CUGUCCAA 1525 TTGGACAG GGCTAGCTACAACGA TCCTCTAG 4329
1712 GACAGGAG G UUUACAGA 1526 TCTGTAAA GGCTAGCTACAACGA CTCCTGTC 4330
1738 CAGAUCAA G UUCAGAAC 1527 GTTCTGAA GGCTAGCTACAACGA TTGATCTG 4331
1751 GAACAAUG G CCUCAUUG 1528 CAATGAGG GGCTAGCTACAACGA CATTGTTC 4332
1771 CUUUUGGG G CCCUUUCA 1529 TGAAAGGG GGCTAGCTACAACGA CCCAAAAG 4333
1792 GAAAUGGA G cuσucucu 1530 AGAGACAG GGCTAGCTACAACGA TCCATTTC 4334
1803 GUCUCUCA G CGCUCCAU 1531 ATGGAGCG GGCTAGCTACAACGA TGAGAGAC 4335
1815 UCCAUCCA G CUUGAGAG 1532 CTCTCAAG GGCTAGCTACAACGA TGGATGGA 4336
1823 GCUUGAGA G UAAGGGAU 1533 ATCCCTTA GGCTAGCTACAACGA TCTCAAGC 4337
1847 CCAGAACA G CCAGUGGA 1534 TCCACTGG GGCTAGCTACAACGA TGTTCTGG 4338
1851 AACAGCCA G UGGAUGAA 1535 TTCATCCA GGCTAGCTACAACGA TGGCTGTT 4339
1862 GAUGAAUG G CACAGUGA 1536 TCACTGTG GGCTAGCTACAACGA CATTCATC 4340
1867 AUGGCACA G UGAUCGUG 1537 CACGATCA GGCTAGCTACAACGA TGTGCCAT 4341
1873 CAGUGAUC G UGGACAGC 1538 GCTGTCCA GGCTAGCTACAACGA GATCACTG 4342
1880 CGUGGACA G CACCGUGG 1539 CCACGGTG GGCTAGCTACAACGA TGTCCACG 4343
1885 ACAGCACC G UGGGAAAG 1540 CTTTCCCA GGCTAGCTACAACGA GGTGCTGT 4344
1926 ACAACGCA G CCUCCCCA 1541 TGGGGAGG GGCTAGCTACAACGA TGCGTTGT 4345
1955 GGAUCCCA G UGGACAGA 1542 TCTGTCCA GGCTAGCTACAACGA TGGGATCC 4346
1965 GGACAGAA G CAAGGUGG 1543 CCACCTTG GGCTAGCTACAACGA TTCTGTCC 4347
1970 GAAGCAAG G UGGCUUUG 1544 CAAAGCCA GGCTAGCTACAACGA CTTGCTTC 4348
1973 GCAAGGUG G CUUUGUAG 1545 CTACAAAG GGCTAGCTACAACGA CACCTTGC 4349
1981 GCUUUGUA G UGGACAAA 1546 TTTGTCCA GGCTAGCTACAACGA TACAAAGC 4350
2002 CCAAAAUG G CCUACCUC 1547 GAGGTAGG GGCTAGCTACAACGA CATTTTGG 4351
2021 AAUCCCAG G CAUUGCUA 1548 TAGCAATG GGCTAGCTACAACGA CTGGGATT 4352
2032 UUGCUAAG G UUGGCACU 1549 AGTGCCAA GGCTAGCTACAACGA CTTAGCAA 4353
2036 UAAGGUUG G CACUUGGA 1550 TCCAAGTG GGCTAGCTACAACGA CAACCTTA 4354
2051 GAAAUACA G UCUGCAAG 1551 CTTGCAGA GGCTAGCTACAACGA TGTATTTC 4355
2059 GUCUGCAA G CAAGCUCA 1552 TGAGCTTG GGCTAGCTACAACGA TTGCAGAC 4356
2063 GCAAGCAA G CUCACAAA 1553 TTTGTGAG GGCTAGCTACAACGA TTGCTTGC 4357
2091 ACUGUCAC G UCCCGUGC 1554 GCACGGGA GGCTAGCTACAACGA GTGACAGT 4358 2096 CACGUCCC G UGCGUCCA 1555 TGGACGCA GGCTAGCTACAACGA GGGACGTG 4359
2100 UCCCGUGC G UCCAAUGC 1556 GCATTGGA GGCTAGCTACAACGA GCACGGGA 4360
2128 CAAUUACA G UGACUUCC 1557 GGAAGTCA GGCTAGCTACAACGA TGTAATTG 4361
2156 GGACACCA G CAAAUUCC 1558 GGAATTTG GGCTAGCTACAACGA TGGTGTCC 4362
2168 AUUCCCCA G CCCUCUGG 1559 CCAGAGGG GGCTAGCTACAACGA TGGGGAAT 4363
2176 GCCCUCUG G UAGUUUAU 1560 ATAAACTA GGCTAGCTACAACGA CAGAGGGC 4364
2179 CUCUGGUA G UUUAUGCA 1561 TGCATAAA GGCTAGCTACAACGA TACCAGAG 4365
2203 GCCAAGGA G CCUCCCCA 1562 TGGGGAGG GGCTAGCTACAACGA TCCTTGGC 4366
2221 UUCUCAGG G CCAGUGUC 1563 GACACTGG GGCTAGCTACAACGA CCTGAGAA 4367
2225 CAGGGCCA G UGUCACAG 1564 CTGTGACA GGCTAGCTACAACGA TGGCCCTG 4368
2233 GUGUCACA G CCCUGAUU 1565 AATCAGGG GGCTAGCTACAACGA TGTGACAC 4369
2248 UUGAAUCA G UGAAUGGA 1566 TCCATTCA GGCTAGCTACAACGA TGATTCAA 4370
2263 GAAAAACA G UUACCUUG 1567 CAAGGTAA GGCTAGCTACAACGA TGTTTTTC 4371
2290 AUAAUGGA G CAGGUGCU 1568 AGCACCTG GGCTAGCTACAACGA TCCATTAT 4372
2294 UGGAGCAG G UGCUGAUG 1569 CATCAGCA GGCTAGCTACAACGA CTGCTCCA 4373
2318 GGAUGACG G UGUCUACU 1570 AGTAGACA GGCTAGCTACAACGA CGTCATCC 4374
2331 UACUCAAG G UAUUUCAC 1571 GTGAAATA GGCTAGCTACAACGA CTTGAGTA 4375
2357 CACGAAUG G UAGAUACA 1572 TGTATCTA GGCTAGCTACAACGA CATTCGTG 4376
2366 UAGAUACA G UGUAAAAG 1573 CTTTTACA GGCTAGCTACAACGA TGTATCTA 4377
2374 GUGUAAAA G UGCGGGCU 1574 AGCCCGCA GGCTAGCTACAACGA TTTTACAC 4378
2380 AAGUGCGG G CUCUGGGA 1575 TCCCAGAG GGCTAGCTACAACGA CCGCACTT 4379
2392 UGGGAGGA G UUAACGCA 1576 TGCGTTAA GGCTAGCTACAACGA TCCTCCCA 4380
2401 UUAACGCA G CCAGACGG 1577 CCGTCTGG GGCTAGCTACAACGA TGCGTTAA 4381
2413 GACGGAGA G UGAUACCC 1578 GGGTATCA GGCTAGCTACAACGA TCTCCGTC 4382
2424 AUACCCCA G CAGAGUGG 1579 CCACTCTG GGCTAGCTACAACGA TGGGGTAT 4383
2429 CCAGCAGA G UGGAGCAC 1580 GTGCTCCA GGCTAGCTACAACGA TCTGCTGG 4384
2434 AGAGUGGA G- CACUGUAC 1581 GTACAGTG GGCTAGCTACAACGA TCCACTCT 4385
2450 CAUACCUG G CUGGAUUG 1582 CAATCCAG GGCTAGCTACAACGA CAGGTATG 4386
2523 CAACACAA G CAAGUGUG 1583 CACACTTG GGCTAGCTACAACGA TTGTGTTG 4387
2527 ACAAGCAA G UGUGUUUC 1584 GAAACACA GGCTAGCTACAACGA TTGCTTGT 4388
2537 GUGUUUCA G CAGAACAU 1585 ATGTTGTG GGCTAGCTACAACGA TGAAACAC 4389
2555 CUCGGGAG G CUCAUUUG 1586 CAAATGAG GGCTAGCTACAACGA CTCCCGAG 4390
2566 CAUUUGUG G CUUCUGAU 1587 ATCAGAAG GGCTAGCTACAACGA CACAAATG 4391
2612 CCCACCUG G CCAAAUCA 1588 TGATTTGG GGCTAGCTACAACGA CAGGTGGG 4392
2632 ACCUGAAG G CGGAAAUU 1589 AATTTCCG GGCTAGCTACAACGA CTTCAGGT 4393
2648 UCACGGGG G CAGUCUCA 1590 TGAGACTG GGCTAGCTACAACGA CCCCGTGA 4394
2651 CGGGGGCA G UCUCAUUA 1591 TAATGAGA GGCTAGCTACAACGA TGCCCCCG 4395
2674 CUUGGACA G CUCCUGGG 1592 CCCAGGAG GGCTAGCTACAACGA TGTCCAAG 4396
2704 AUGGAACA G CUCACAAG 1593 CTTGTGAG GGCTAGCTACAACGA TGTTCCAT 4397
2712 GCUCACAA G UAUAUCAU 1594 ATGATATA GGCTAGCTACAACGA TTGTGAGC 4398
2729 UCGAAUAA G UACAAGUA 1595 TACTTGTA GGCTAGCTACAACGA TTATTCGA 4399
2735 AAGUACAA G UAUUCUUG 1596 CAAGAATA GGCTAGCTACAACGA TTGTACTT 4400
2757 AGAGACAA G UUCAAUGA 1597 TCATTGAA GGCTAGCTACAACGA TTGTCTCT 4401
2776 CUCUUCAA G UGAAUACU 1598 AGTATTCA GGCTAGCTACAACGA TTGAAGAG 4402
2806 CAAAGGAA G CCAACUCU 1599 AGAGTTGG GGCTAGCTACAACGA TTCCTTTG 4403
2821 CUGAGGAA G UCUUUUUG 1600 CAAAAAGA GGCTAGCTACAACGA TTCCTCAG 4404
2861 UGAAAAUG G CACAGAUC 1601 GATCTGTG GGCTAGCTACAACGA CATTTTCA 4405
2887 CUAUUCAG G CUGUUGAU 1602 ATCAACAG GGCTAGCTACAACGA CTGAATAG 4406
2899 UUGAUAAG G UCGAUCUG 1603 CAGATCGA GGCTAGCTACAACGA CTTATCAA 4407
2935 UUGCACGA G UAUCUUUG 1604 CAAAGATA GGCTAGCTACAACGA TCGTGCAA 4408
2978 GACACCUA G UCCUGAUG 1605 CATCAGGA GGCTAGCTACAACGA TAGGTGTC 4409
2991 GAUGAAAC G UCUGCUCC 1606 GGAGCAGA GGCTAGCTACAACGA GTTTCATC 4410
3023 UAUCAACA G CACCAUUC 1607 GAATGGTG GGCTAGCTACAACGA TGTTGATA 4411
3035 CAUUCCUG G CAUUCACA 1608 TGTGAATG GGCTAGCTACAACGA CAGGAATG 4412
3063 AUGUGGAA G UGGAUAGG 1609 CCTATCCA GGCTAGCTACAACGA TTCCACAT 4413
3081 GAACUGCA G CUGUCAAU 1610 ATTGACAG GGCTAGCTACAACGA TGCAGTTC 4414 3091 UGUCAAUA G CCUAGGGC 1611 GCCCTAGG GGCTAGCTACAACGA TATTGACA 4415
3098 AGCCUAGG G CUGAAUUU 1612 AAATTCAG GGCTAGCTACAACGA CCTAGGCT 4416
3189 UGUAGGGG G CGAUAUAC 1613 GTATATCG GGCTAGCTACAACGA CCCCTACA 4417
3242 UGUAGGGG G CGAUAUAC 1613 GTATATCG GGCTAGCTACAACGA CCCCTACA 4417
3210 UGUAUAUA G UACAUUUA 1614 TAAATGTA GGCTAGCTACAACGA TATATACA 4418
3279 UGUAGGGG G CGAUAAAA 1615 TTTTATCG GGCTAGCTACAACGA CCCCTACA 4419
21 UGGUACAA A UGGAUGUG 1616 CACATCCA GGCTAGCTACAACGA TTGTACCA 4420
25 ACAAAUGG A UGUGGAAU 1617 ATTCCACA GGCTAGCTACAACGA CCATTTGT 4421
32 GAUGUGGA A UAUAAUUG 1618 CAATTATA GGCTAGCTACAACGA TCCACATC 4422
37 GGAAUAUA A UUGAAUAU 1619 ATATTCAA GGCTAGCTACAACGA TATATTCC 4423
42 AUAAUUGA A UAUUUUCU 1620 AGAAAATA GGCTAGCTACAACGA TCAATTAT 4424
114 GAAGGCAG A UGGAAAUA 1621 TATTTCCA GGCTAGCTACAACGA CTGCCTTC 4425
120 AGAUGGAA A UAUUUACA 1622 TGTAAATA GGCTAGCTACAACGA TTCCATCT 4426
137 AGUACGCA A UUUGAGAC 1623 GTCTCAAA GGCTAGCTACAACGA TGCGTACT 4427
144 AAUUUGAG A CUAAGAUA 1624 TATCTTAG GGCTAGCTACAACGA CTCAAATT 4428
150 AGACUAAG A UAUUGUUA 1625 TAACAATA GGCTAGCTACAACGA CTTAGTCT 4429
176 UAUUGAAG A CAAGAGCA 1626 TGCTCTTG GGCTAGCTACAACGA CTTCAATA 4430
185 CAAGAGCA A UAGUAAAA 1627 TTTTACTA GGCTAGCTACAACGA TGCTCTTG 4431
193 AUAGUAAA A CACAUCAG 1628 CTGATGTG GGCTAGCTACAACGA TTTACTAT 4432
217 GGUUAAAG A CCUGUGAU 1629 ATCACAGG GGCTAGCTACAACGA CTTTAACC 4433
224 GACCUGUG A UAAACCAC 1630 GTGGTTTA GGCTAGCTACAACGA CACAGGTC 4434
228 UGUGAUAA A CCACUUCC 1631 GGAAGTGG GGCTAGCTACAACGA TTATCACA 4435
238 CACUUCCG A UAAGUUGG 1632 CCAACTTA GGCTAGCTACAACGA CGGAAGTG 4436
249 AGUUGGAA A CGUGUGUC 1633 GACACACG GGCTAGCTACAACGA TTCCAACT 4437
284 UAUAUAUA A UGGUAAAG 1634 CTTTACCA GGCTAGCTACAACGA TATATATA 4438
297 AAAGAAAG A CACCUUCG 1635 CGAAGGTG GGCTAGCTACAACGA CTTTCTTT 4439
308 CCUUCGUA A CCCGCAUU 1636 AATGCGGG GGCTAGCTACAACGA TACGAAGG 4440
331 AGAGAGGA A UCACAGGG 1637 CCCTGTGA GGCTAGCTACAACGA TCCTCTCT 4441
342 ACAGGGAG A UGUACAGC 1638 GCTGTACA GGCTAGCTACAACGA CTCCCTGT 4442
352 GUACAGCA A UGGGGCCA 1639 TGGCCCCA GGCTAGCTACAACGA TGCTGTAC 4443
385 UCAUCUUG A UUCUUCAC 1640 GTGAAGAA GGCTAGCTACAACGA CAAGATGA 4444
416 CCUGAGUA A UUCACUCA 1641 TGAGTGAA GGCTAGCTACAACGA TACTCAGG 4445
434 UCAGCUGA A CAACAAUG 1642 CATTGTTG GGCTAGCTACAACGA TCAGCTGA 4446
437 GCUGAACA A CAAUGGCU 1643 AGCCATTG GGCTAGCTACAACGA TGTTCAGC 4447
440 GAACAACA A UGGCUAUG ■ 1644 CATAGCCA GGCTAGCTACAACGA TGTTGTTC 4448
466 UCGUUGCA A UCGACCCC 1645 GGGGTCGA GGCTAGCTACAACGA TGCAACGA 4449
470 UGCAAUCG A CCCCAAUG 1646 CATTGGGG GGCTAGCTACAACGA CGATTGCA 4450
476 CGACCCCA A UGUGCCAG 1647 CTGGCACA GGCTAGCTACAACGA TGGGGTCG 4451
488 GCCAGAAG A UGAAACAC 1648 GTGTTTCA GGCTAGCTACAACGA CTTCTGGC 4452
493 AAGAUGAA A CACUCAUU 1649 AATGAGTG GGCTAGCTACAACGA TTCATCTT 4453
.504 CUCAUUCA A CAAAUAAA 1650 TTTATTTG GGCTAGCTACAACGA TGAATGAG 4454
508 UUCAACAA A UAAAGGAC 1651 GTCCTTTA GGCTAGCTACAACGA TTGTTGAA 4455
515 AAUAAAGG A CAUGGUGA 1652 TCACCATG GGCTAGCTACAACGA CCTTTATT 4456
523 ACAUGGUG A CCCAGGCA 1653 TGCCTGGG GGCTAGCTACAACGA CACCATGT 4457
564 GGAAAGCG A UUUUAUUU 1654 AAATAAAA GGCTAGCTACAACGA CGCTTTCC 4458
578 UUUCAAAA A UGUUGCCA 1655 TGGCAACA GGCTAGCTACAACGA TTTTGAAA 4459
592 CCAUUUUG A UUCCUGAA 1656 TTCAGGAA GGCTAGCTACAACGA CAAAATGG 4460
601 UUCCUGAA A CAUGGAAG 1657 CTTCCATG GGCTAGCTACAACGA TTCAGGAA 4461
610 CAUGGAAG A CAAAGGCU 1658 AGCCTTTG GGCTAGCTACAACGA CTTCCATG 4462
620 AAAGGCUG A CUAUGUGA 1659 TCACATAG GGCTAGCTACAACGA CAGCCTTT 4463
630 UAUGUGAG A CCAAAACU 1660 AGTTTTGG GGCTAGCTACAACGA CTCACATA 4464
636 AGACCAAA A CUUGAGAC 1661 GTCTCAAG GGCTAGCTACAACGA TTTGGTCT 4465
643 AACUUGAG A CCUACAAA 1662 TTTGTAGG GGCTAGCTACAACGA CTCAAGTT 4466
653 CUACAAAA A UGCUGAUG 1663 CATCAGCA GGCTAGCTACAACGA TTTTGTAG 4467
659 AAAUGCUG A UGUUCUGG 1664 CCAGAACA GGCTAGCTACAACGA CAGCATTT 4468
692 UCCAGGUA A UGAUGAAC 1665 GTTCATCA GGCTAGCTACAACGA TACCTGGA 4469
Figure imgf000113_0001
1425 CAAAGUGA A CUCAUACA 1722 TGTATGAG GGCTAGCTACAACGA TCACTTTG 4526
1435 UCAUACAG A UAAACAGU 1723 ACTGTTTA GGCTAGCTACAACGA CTGTATGA 4527
1439 ACAGAUAA A CAGUGGCA 1724 TGCCACTG GGCTAGCTACAACGA TTATCTGT 4528
1451 UGGCAGUG A CAGGGACA 1725 TGTCCCTG GGCTAGCTACAACGA CACTGCCA 4529
1457 UGACAGGG A CACACUCG 1726 CGAGTGTG GGCTAGCTACAACGA CCCTGTCA 4530
1473 GCCAAAAG A UUACCUGC 1727 GCAGGTAA GGCTAGCTACAACGA CTTTTGGC 4531
1498 CAGGAGGG A CGUCCAUC 1728 GATGGACG GGCTAGCTACAACGA CCCTCCTG 4532
1521 GGGCUUCG A UCGGCAUU 1729 AATGCCGA GGCTAGCTACAACGA CGAAGCCC 4533
1537 UUACUGUG A UUAGGAAG 1730 CTTCCTAA GGCTAGCTACAACGA CACAGTAA 4534
1548 AGGAAGAA A UAUCCAAC 1731 GTTGGATA GGCTAGCTACAACGA TTCTTCCT 4535
1555 AAUAUCCA A CUGAUGGA 1732 TCCATCAG GGCTAGCTACAACGA TGGATATT 4536
1559 UCCAACUG A UGGAUCUG 1733 CAGATCCA GGCTAGCTACAACGA CAGTTGGA ■ 4537
1563 ACUGAUGG A UCUGAAAU 1734 ATTTCAGA GGCTAGCTACAACGA CCATCAGT 4538
1570 GAUCUGAA A UUGUGCUG 1735 CAGCACAA GGCTAGCTACAACGA TTCAGATC 4539
1582 UGCUGCUG A CGGAUGGG 1736 CCCATCCG GGCTAGCTACAACGA CAGCAGCA 4540
1586 GCUGACGG A UGGGGAAG 1737 CTTCCCCA GGCTAGCTACAACGA CCGTCAGC 4541
1595 UGGGGAAG A CAACACUA 1738 TAGTGTTG GGCTAGCTACAACGA CTTCCCCA 4542
1598 GGAAGACA A CACUAUAA 1739 TTATAGTG GGCTAGCTACAACGA TGTCTTCC 4543
1619 GUGCUUUA A CGAGGUCA 1740 TGACCTCG GGCTAGCTACAACGA TAAAGCAC 4544
1629 GAGGUCAA A CAAAGUGG 1741 CCACTTTG GGCTAGCTACAACGA TTGACCTC 4545
1683 GCUCAAGA A CUAGAGGA 1742 TCCTCTAG GGCTAGCTACAACGA TCTTGAGC 4546
1702 UGUCCAAA A UGACAGGA 1743 TCCTGTCA GGCTAGCTACAACGA TTTGGACA 4547
1705 CCAAAAUG A CAGGAGGU 1744 ACCTCCTG GGCTAGCTACAACGA CATTTTGG 4548
1720 GUUUACAG A CAUAUGCU 1745 AGCATATG GGCTAGCTACAACGA CTGTAAAC 4549
1733 UGCUUCAG A UCAAGUUC 1746 GAACTTGA GGCTAGCTACAACGA CTGAAGCA 4550
1745 AGUUCAGA A CAAUGGCC 1747 GGCCATTG GGCTAGCTACAACGA TCTGAACT 4551
1748 UCAGAACA A UGGCCUCA 1748 TGAGGCCA GGCTAGCTACAACGA TGTTCTGA 4552
1760 CCUCAUUG A UGCUUUUG 1749 CAAAAGCA GGCTAGCTACAACGA CAATGAGG 4553
1787 AUCAGGAA A UGGAGCUG 1750 CAGCTCCA GGCTAGCTACAACGA TTCCTGAT 4554
1830 AGUAAGGG A UUAACCCU 1751 AGGGTTAA GGCTAGCTACAACGA CCCTTACT 4555
1834 AGGGAUUA A CCCUCCAG 1752 CTGGAGGG GGCTAGCTACAACGA TAATCCCT 4556
1844 CCUCCAGA A CAGCCAGU 1753 ACTGGCTG GGCTAGCTACAACGA TCTGGAGG 4557
1855 GCCAGUGG A UGAAUGGC 1754 GCCATTCA GGCTAGCTACAACGA CCACTGGC 4558
1859 GUGGAUGA A UGGCACAG 1755 CTGTGCCA GGCTAGCTACAACGA TCATCCAC 4559
1870 GCACAGUG A UCGUGGAC 1756 GTCCACGA GGCTAGCTACAACGA CACTGTGC 4560
1877 GAUCGUGG A CAGCACCG 1757 CGGTGCTG GGCTAGCTACAACGA CCACGATC 4561
1895 GGGAAAGG A CACUUUGU 1758 ACAAAGTG GGCTAGCTACAACGA CCTTTCCC 4562
1918 UCACCUGG A CAACGCAG 1759 CTGCGTTG GGCTAGCTACAACGA CCAGGTGA 4563
1921 CCUGGACA A CGCAGCCU 1760 AGGCTGCG GGCTAGCTACAACGA TGTCCAGG 4564
1936 CUCCCCAA A UCCUUCUC 1761 GAGAAGGA GGCTAGCTACAACGA TTGGGGAG 4565
1949 UCUCUGGG A UCCCAGUG 1762 CACTGGGA GGCTAGCTACAACGA CCCAGAGA 4566
1959 CCCAGUGG A CAGAAGCA 1763 TGCTTCTG GGCTAGCTACAACGA CCACTGGG 4567
1985 UGUAGUGG A CAAAAACA 1764 TGTTTTTG GGCTAGCTACAACGA CCACTACA 4568
1991 GGACAAAA A CACCAAAA 1765 TTTTGGTG GGCTAGCTACAACGA TTTTGTCC 4569
1999 ACACCAAA A UGGCCUAC 1766 GTAGGCCA GGCTAGCTACAACGA TTTGGTGT 4570
2014 ACCUCCAA A UCCCAGGC 1767 GCCTGGGA GGCTAGCTACAACGA TTGGAGGT 4571
2046 ACUUGGAA A UACAGUCU 1768 AGACTGTA GGCTAGCTACAACGA TTCCAAGT 4572
2071 GCUCACAA A CCUUGACC 1769 GGTCAAGG GGCTAGCTACAACGA TTGTGAGC 4573
2077 AAACCUUG A CCCUGACU 1770 AGTCAGGG GGCTAGCTACAACGA CAAGGTTT 4574
2083 UGACCCUG A CUGUCACG 1771 CGTGACAG GGCTAGCTACAACGA CAGGGTCA 4575
2105 UGCGUCCA A UGCUACCC 1772 GGGTAGCA GGCTAGCTACAACGA TGGACGCA 4576
2122 UGCCUCCA A UUACAGUG 1773 CACTGTAA GGCTAGCTACAACGA TGGAGGCA 4577
2131 UUACAGUG A CUUCCAAA 1774 TTTGGAAG GGCTAGCTACAACGA CACTGTAA 4578
2140 CUUCCAAA A CGAACAAG 1775 CTTGTTCG GGCTAGCTACAACGA TTTGGAAG 4579
2144 CAAAACGA A CAAGGACA 1776 TGTCCTTG GGCTAGCTACAACGA TCGTTTTG 4580
2150 GAACAAGG A CACCAGCA 1777 TGCTGGTG GGCTAGCTACAACGA CCTTGTTC 4581
Figure imgf000115_0001
2867 UGGCACAG A UCUUUUCA 1834 TGAAAAGA GGCTAGCTACAACGA CTGTGCCA 4638
2894 GGCUGUUG A UAAGGUCG 1835 CGACCTTA GGCTAGCTACAACGA CAACAGCC 4639
2903 UAAGGUCG A UCUGAAAU 1836 ATTTCAGA GGCTAGCTACAACGA CGACCTTA 4640
2910 GAUCUGAA A UCAGAAAU 1837 ATTTCTGA GGCTAGCTACAACGA TTCAGATC 4641
2917 AAUCAGAA A UAUCCAAC 1838 GTTGGATA GGCTAGCTACAACGA TTCTGATT 4642
2924 AAUAUCCA A CAUUGCAC 1839 GTGCAATG GGCTAGCTACAACGA TGGATATT 4643
2959 CUCCACAG A CUCCGCCA 1840 TGGCGGAG GGCTAGCTACAACGA CTGTGGAG 4644
2971 CGCCAGAG A CACCUAGU 1841 ACTAGGTG GGCTAGCTACAACGA CTCTGGCG 4645
2984 UAGUCCUG A UGAAACGU 1842 ACGTTTCA GGCTAGCTACAACGA CAGGACTA 4646
2989 CUGAUGAA A CGUCUGCU 1843 AGCAGACG GGCTAGCTACAACGA TTCATCAG 4647
3008 UUGUCCUA A UAUUCAUA 1844 TATGAATA GGCTAGCTACAACGA TAGGACAA 4648
3020 UCAUAUCA A CAGCACCA 1845 TGGTGCTG GGCTAGCTACAACGA TGATATGA 4649
3052 UUUUAAAA A UUAUGUGG 1846 CCACATAA GGCTAGCTACAACGA TTTTAAAA 4650
3067 GGAAGUGG A UAGGAGAA 1847 TTCTCCTA GGCTAGCTACAACGA CCACTTCC 4651
3075 AUAGGAGA A CUGCAGCU 1848 AGCTGCAG GGCTAGCTACAACGA TCTCCTAT 4652
3088 AGCUGUCA A UAGCCUAG 1849 CTAGGCTA GGCTAGCTACAACGA TGACAGCT 4653
3103 AGGGCUGA A UUUUUGUC 1850 GACAAAAA GGCTAGCTACAACGA TCAGCCCT 4654
3114 UUUGUCAG A UAAAUAAA 1851 TTTATTTA GGCTAGCTACAACGA CTGACAAA 4655
3118 UCAGAUAA A UAAAAUAA 1852 TTATTTTA GGCTAGCTACAACGA TTATCTGA 4656
3123 UAAAUAAA A UAAAUCAU 1853 ATGATTTA GGCTAGCTACAACGA TTTATTTA 4657
3127 UAAAAUAA A UCAUUCAU 1854 ATGAATGA GGCTAGCTACAACGA TTATTTTA 4658
3146 UUUUUUUG A UUAUAAAA 1855 TTTTATAA GGCTAGCTACAACGA CAAAAAAA 4659
3154 AUUAUAAA A UUUUCUAA 1856 TTAGAAAA GGCTAGCTACAACGA TTTATAAT 4660
3164 UUUCUAAA A UGUAUUUU 1857 AAAATACA GGCTAGCTACAACGA TTTAGAAA 4661
3175 UAUUUUAG A CUUCCUGU 1858 ACAGGAAG GGCTAGCTACAACGA CTAAAATA 4662
3265 UAUUUUAG A CUUCCUGU 1858 ACAGGAAG GGCTAGCTACAACGA CTAAAATA 4662
3192 AGGGGGCG A UAUACUAA 1859 TTAGTATA GGCTAGCTACAACGA CGCCCCCT 4663
3245 AGGGGGCG A UAUACUAA 1859 TTAGTATA GGCTAGCTACAACGA CGCCCCCT 4663
3201 UAUACUAA A UGUAUAUA 1860 TATATACA GGCTAGCTACAACGA TTAGTATA 4664
3225 UAUACUAA A UGUAUUCC 1861 GGAATACA GGCTAGCTACAACGA TTAGTATA 4665
3254 UAUACUAA A UGUAUUUU 1862 AAAATACA GGCTAGCTACAACGA TTAGTATA 4666
3282 AGGGGGCG A UAAAAUAA 1863 TTATTTTA GGCTAGCTACAACGA CGCCCCCT 4667
3287 GCGAUAAA A UAAAAUGC 1864 GCATTTTA GGCTAGCTACAACGA TTTATCGC 4668
3292 AAAAUAAA A UGCUAAAC 1865 GTTTAGCA GGCTAGCTACAACGA TTTATTTT 4669
3299 AAUGCUAA A CAACUGGG 1866 CCCAGTTG GGCTAGCTACAACGA TTAGCATT 4670
3302 GCUAAACA A CUGGGUAA 1867 TTACCCAG GGCTAGCTACAACGA TGTTTAGC 4671
Input Sequence = NM_001285. Cut Site = R/Y
Arm Length = 8. Core Sequence = GGCTAGCTACAACGA
NM_001285 (Homo sapiens chloride channel, calcium activated, 1 (CLCAl) mRNA, 3311 bp) Table VIII: Human CLCAl Amberzyme and Target Sequence 249.021
Pos Substrate Seq Amberzyme Rz
ID Seq
No. ID No.
40 AUAUAAUU G AAUAUUUU 1211 AAAAUAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUUAUAU 4672
67 GGGAGCAU G AAGAGGUG 1212 CACCUCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUGCUCCC 4673
78 GAGGUGUϋ G AGGUUAUG 1213 CAUAACCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AACACCUC 4674
106 GGACAGCU G AAGGCAGA 1214 UCUGCCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCUGUGC 4675
134 ACAAGUAC G CAAUUUGA 1215 UCAAAUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUACUUGU 4676
141 CGCAAUUU G AGACUAAG 1216 CUUAGUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAUUGCG 4677
172 CUCCUAUU G AAGACAAG 1217 CUUGUCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUAGGAG 4678
223 AGACCUGU G AUAAACCA 1218 UGGUUUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAGGUCU 4679
237 CCACUUCC G AUAAGUUG 1219 CAACUUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GGAAGUGG 4680
312 CGUAACCC G CAUUUUCC 1220 GGAAAAUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GGGUUACG 4681
384 UUCAUCUU G AUUCUUCA 1221 UGAAGAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAGAUGAA 4682
411 GGGGCCCU G AGUAAUUC 1222 GAAUUACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGGCCCC 4683
432 AUUCAGCU G AACAACAA 1223 UUGUUGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCUGAAU 4684
448 AUGGCUAU G AAGGCAUU 1224 AAUGCCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUAGCCAU 4685
463 UUGUCGUU G CAAUCGAC 1225 GUCGAUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AACGACAA 4686
469 UUGCAAUC G ACCCCAAU 1226 AUUGGGGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GAUUGCAA 4687
480 CCCAAUGU G CCAGAAGA 1227 UCUUCUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAUUGGG 4688
490 CAGAAGAU G AAACACUC 1228 GAGUGUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCUUCUG 4689
522 GACAUGGU G ACCCAGGC 1229 GCCUGGGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACCAUGUC 4690
547 AUCUGUUU G AAGCUACA 1230 UGUAGCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAACAGAU 4691
563 AGGAAAGC G AUUUUAUU 1231 AAUAAAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GCUUUCCU 4692
583 AAAAUGUU G CCAUUUUG 1232 CAAAAUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AACAUUUU 4693
591 GCCAUUUU G AUUCCUGA 1233 UCAGGAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAAUGGC 4694
598 UGAUUCCU G AAACAUGG 1234 CCAUGUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGAAUCA 4695
619 CAAAGGCU G ACUAUGUG 1235 CACAUAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCCUUUG 4696
627 GACUAUGU G AGACCAAA 1236 UUUGGUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAUAGUC 4697
640 CAAAACUU G AGACCUAC 1237 GUAGGUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAGUUUUG 4698
655 ACAAAAAU G CUGAUGUU 1238 AACAUCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUUUGU 4699
658 AAAAUGCU G AUGUUCUG 1239 CAGAACAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCAUUUU 4700
670 UUCUGGUU G CUGAGUCU 1240 AGACUCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AACCAGAA 4701
673 UGGUUGCU G AGUCUACU 1241 AGUAGACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCAACCA 4702
694 CAGGUAAU G AUGAACCC 1242 GGGUUCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUACCUG 4703
697 . GUAAUGAU G AACCCUAC 1243 GUAGGGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCAUUAC 4704
709 CCUACACU G AGCAGAUG 1244 CAUCUGCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUGUAGG 4705
739 AGAAGGGU G AAAGGAUC 1245 GAUGCUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACCCUUCU 4706
760 UCACUCCU G AUUUCAUU 1246 AAUGAAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGAGUGA 4707
769 AUUUCAUU G CAGGAAAA 1247 UUUUCCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUGAAAU 4708
787 AGUUAGCU G AAUAUGGA 1248 UCCAUAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCUAACU 4709
820 UUGUCCAU G AGUGGGCU 1249 AGCCCACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUGGACAA 4710
836 UCAUCUAC G AUGGGGAG 1250 CUCCCCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUAGAUGA 4711
850 GAGUAUUU G ACGAGUAC 1251 GUACUCGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAUACUC 4712
853 UAUUUGAC G AGUACAAU 1252 AUUGUACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUCAAAUA 4713
865 ACAAUAAU G AUGAGAAA 1253 UUUCUCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUAUUGU 4714
868 AUAAUGAU G AGAAAUUC 1254 GAAUUUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCAUUAU 4715
980 CAAAAGAU G CACAUUCA 1255 UGAAUGUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCUUUUG 4716
1009 GACUCUAU G AAAAAGGA 1256 UCCUUUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUAGAGUC 4717
1021 AAGGAUGU G AGUUUGUU 1257 AACAAACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAUCCUU 4718
1040 CCAAUCCC G CCAGACGG 1258 CCGUCUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GGGAUUGG 4719
1069 UAAUGUUU G CACAACAU 1259 AUGUUGUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAACAUUA 4720
1081 AACAUGUU G AUUCUAUA 1260 UAUAGAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AACAUGUU 4721
1093 CUAUAGUU G AAUUCUGU 1261 ACAGAAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AACUAUAG 4722
1151 UCAAAAAU G CAAUCUCC 1262 GGAGAUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUUUGA 4723
1160 CAAUCUCC G AAGCACAU 1263 AUGUGCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GGAGAUUG 4724
1176 UGGGAAGU G AUCCGUGA 1254 UCACGGAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUUCCCA 4725
1183 UGAUCCGU G AUUCUGAG 1265 CUCAGAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACGGAUCA 4726
1189 GUGAUUCU G AGGACUUU 1266 AAAGUCCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAAUCAC 4727
1215 ACUCCUAU G ACAACACA 1267 UGUGUUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUAGGAGU 4728
1248 UUCUCAUU G CUGCAGAU 1268 AUCUGCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUGAGAA 4729
1251 UCAUUGCU G CAGAUUGG 1269 CCAAUCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCAAUGA 4730
1285 UAGUCCUU G ACAAAUCU 1270 AGAUUUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAGGACUA 4731
1305 AGCAUGGC G ACUGGUAA 1271 UUACCAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GCCAUGCU 4732
1316 UGGUAACC G CCUCAAUC 1272 GAUUGAGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GGUUACCA 4733
1325 CCUCAAUC G ACUGAAUC 1273 GAUUCAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GAUUGAGG 4734
1329 AAUCGACU G AAUCAAGC 1274 GCUUGAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUCGAUU 4735
1353 CUUUUCCU G CUGCAGAC 1275 GUCUGCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGAAAAG 4736
1356 UUCCUGCU G CAGACAGU 1276 ACUGUCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCAGGAA 4737
1366 AGACAGUU G AGCUGGGG 1277 CCCCAGCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AACUGUCU 4738
1392 GGGAUGGU G ACAUUUGA 1278 UCAAAUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACCAUCCC 4739
1399 UGACAUUU G ACAGUGCU 1279 AGCACUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAUGUCA 4740
1405 UUGACAGU G CUGCCCAU 1280 AUGGGCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUGUCAA 4741
1408 ACAGUGCU G CCCAUGUA 1281 UACAUGGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCACUGU 4742
1423 UACAAAGU G AACUCAUA 1282 UAUGAGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUUUGUA 4743
1450 GUGGCAGU G ACAGGGAC 1283 GUCCCUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUGCCAC 4744
1465 ACACACUC G CCAAAAGA 1284 UCUUUUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GAGUGUGU 4745
1480 GAUUACCU G CAGCAGCO 1285 AGCUGCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGUAAUC 4746
1508 GUCCAUCU G CAGCGGGC 1286 GCCCGCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAUGGAC 4747
1520 CGGGCUUC G AUCGGCAU 1287 AUGCCGAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GAAGCCCG 4748
1536 UUUACUGU G AUUAGGAA 1288 UUCCUAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAGUAAA 4749
1558 AUCCAACU G AUGGAUCU 1289 AGAUCCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUUGGAU 4750
1567 AUGGAUCU G AAAUUGUG 1290 CACAAUUϋ GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAUCCAU 4751
1575 GAAAUUGU G CUGCUGAC 1291 GUCAGCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAAUUUC 4752
1578 AUUGUGCU G CUGACGGA 1292 UCCGUCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCACAAU 4753
1581 GUGCUGCU G ACGGAUGG 1293 CCAUCCGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCAGCAC 4754
1613 AAGUGGGU G CUUUAACG 1294 CGUUAAAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACCCACUU 4755
1621 GCUUUAAC G AGGUCAAA 1295 UUUGACCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUUAAAGC 4756
1639 AAAGUGGU G CCAUCAUC 1296 GAUGAUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACCACUUU 4757
1657 ACACAGUC G CUUUGGGG 1297 CCCCAAAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GACUGUGU 4758
1672 GGCCCUCU G CAGCUCAA 1298 UUGAGCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAGGGCC 4759
1704 UCCAAAAU G ACAGGAGG 1299 CCUCCUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUUGGA 4760
1726 AGACAUAU G CUUCAGAU 1300 AUCUGAAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUAUGUCU 4761
1759 GCCUCAUU G AUGCUUUϋ 1301 AAAAGCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUGAGGC 4762
1762 UCAUUGAU G CUUUUGGG 1302 CCCAAAAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCAAUGA 4763
1805 CUCUCAGC G CUCCAUCC 1303 GGAUGGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GCUGAGAG 4764
1819 UCCAGCUU G AGAGUAAG 1304 CUUACUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAGCUGGA 4765
1857 CAGUGGAU G AAUGGCAC 1305 GUGCCAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCCACUG 4766
1869 GGCACAGU G AUCGUGGA 1306 UCCACGAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUGUGCC 4767
1923 UGGACAAC G CAGCCUCC 1307 GGAGGCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUUGUCCA 4768
2026 CAGGCAUU G CUAAGGUU 1308 AACCUUAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUGCCUG 4769
2055 UACAGUCU G CAAGCAAG 1309 CUUGCUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGACUGUA 4770
2076 CAAACCUU G ACCCUGAC 1310 GUCAGGGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAGGUUUG 4771
2082 UUGACCCU G ACUGUCAC 1311 GUGACAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGGUCAA 4772
2098 CGUCCCGU G CGUCCAAU 1312 AUUGGACG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACGGGACG 4773
2107 CGUCCAAU G CUACCCUG 1313 CAGGGUAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUGGACG 4774
2115 GCUACCCU G CCUCCAAU 1314 AUUGGAGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGGUAGC 4775
2130 AUUACAGU G ACUUCCAA 1315 UUGGAAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUGUAAU 4776
2142 UCCAAAAC G AACAAGGA 1316 UCCUUGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUUUUGGA 4777
2185 UAGUUUAU G CAAAUAUU 1317 AAUAUUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUAAACUA 4778
2195 AAAUAUUC G CCAAGGAG 1318 CUCCUUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GAAUAUUU 4779
2238 ACAGCCCU G AUUGAAUC 1319 GAUUCAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGGCUGU 4780
2242 CCCUGAUU G AAUCAGUG 1320 CACUGAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUCAGGG 4781
2250 GAAUCAGU G AAUGGAAA 1321 UUUCCAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUGAUUC 4782
2296 GAGCAGGU G CUGAUGCU 1322 AGCAUCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACCUGCUC 4783
2299 CAGGUGCU G AUGCUACU 1323 AGUAGCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCACCUG 4784
2302 GUGCUGAU G CUACUAAG 1324 CUUAGUAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCAGCAC 4785
2314 CUAAGGAU G ACGGUGUC 1325 GACACCGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCCUUAG 4786
2347 CAACUUAU G ACACGAAU 1326 AUUCGUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUAAGUUG 4787
2352 UAUGACAC G AAUGGUAG 1327 CUACCAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUGUCAUA 4788
2376 GUAAAAGU G CGGGCUCU 1328 AGAGCCCG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUUUUAC 4789
2398 GAGUUAAC G CAGCCAGA 1329 UCUGGCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUUAACUC 4790
2415 CGGAGAGU G AUACCCCA 1330 UGGGGUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUCUCCG 4791
2458 GCUGGAUϋ G AGAAUGAU 1331 AUCAUUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUCCAGC 4792
2464 UUGAGAAU G AUGAAAUA 1332 UAUUUCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUCUCAA 4793
2467 AGAAUGAU G AAAUACAA 1333 UUGUAUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCAUUCU 4794
2494 CAAGACCU G AAAUUAAU 1334 AUUAAUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGUCUUG 4795
2509 AUAAGGAU G AUGUUCAA 1335 UUGAACAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCCUUAU 4796
2572 UGGCUUCU G AUGUCCCA 1336 UGGGACAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAAGCCA 4797
2584 UCCCAAAU G CUCCCAUA 1337 UAUGGGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUGGGA 4798
2596 CCAUACCU G AUCUCUUC 1338 GAAGAGAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGUAUGG^ 4799
2623 AAAUCACC G ACCUGAAG 1339 CUUCAGGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GGUGAUUU 4800
2628 ACCGACCU G AAGGCGGA 1340 UCCGCCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGUCGGU 4801
2664 AUUAAUCU G ACUUGGAC 1341 GUCCAAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAUUAAU 4802
2686 CUGGGGAU G AUUAUGAC 1342 GUCAUAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCCCCAG 4803
2692 AUGAUUAU G ACCAUGGA 1343 UCCAUGGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUAAUCAU 4804
2723 UAUCAUUC G AAUAAGUA 1344 UACUUAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GAAUGAUA 4805
2743 GUAUUCUU G AUCUCAGA 1345 UCUGAGAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAGAAUAC 4806
2764 AGUUCAAU G AAUCUCUU 1346 AAGAGAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUGAACU 4807
2778 CUUCAAGU G AAUACUAC 1347 GUAGUAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUUGAAG 4808
2788 AUACUACU G CUCUCAUC 1348 GAUGAGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUAGUAU 4809
2815 CCAACUCU G AGGAAGUC 1349 GACUUCCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAGUUGG 4810
2854 UUACUUUϋ G AAAAUGGC 1350 GCCAUUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAAGUAA 4811
2878 UUUUGAUU G CUAUUCAG 1351 CUGAAUAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUGAAAA 4812
2893 AGGCUGUU G AUAAGGUC 1352 GACCUUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AACAGCCU 4813
2902 AUAAGGUC G AUCUGAAA 1353 UUUCAGAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GACCUUAU 4814
2907 GUCGAUCU G AAAUCAGA 1354 UCUGAUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAUCGAC 4815
2929 CCAACAUU G CACGAGUA 1355 UACUCGUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUGUUGG 4816
2933 CAUUGCAC G AGUAUCUU 1356 AAGAUACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUGCAAUG 4817
2964 CAGACUCC G CCAGAGAC 1357 GUCUCUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GGAGUCUG 4818
2983 CUAGUCCU G AUGAAACG 1358 CGUUUCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGACUAG 4819
2986 GUCCUGAU G AAACGUCU 1359 AGACGUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCAGGAC 4820
2995 AAACGUCU G CUCCUUGU 1360 ACAAGGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGACGUUU 4821
3078 GGAGAACU G CAGCUGUC 1361 GACAGCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUUCUCC 4822
3101 CUAGGGCU G AAUUUUUG 1362 CAAAAAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCCCUAG 4823
3145 CUUUUUUU G AUUAUAAA 1363 UUUAUAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAAAAAG 4824
3191 UAGGGGGC G AUAUACUA 1364 UAGUAUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GCCCCCUA 4825
3244 UAGGGGGC G AUAUACUA 1364 UAGUAUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GCCCCCUA 4825
3281 UAGGGGGC G AUAAAAUA 1365 UAUUUUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GCCCCCUA 4826
3294 AAUAAAAU G CUAAACAA 1366 UUGUUUAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUUAUU 4827
27 AAAUGGAU G UGGAAUAU 1367 AUAUUCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCCAUUU 4828
52 AUUUUCUU G UUUAAGGG 1368 CCCUUAAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAGAAAAU 4829
75 GAAGAGGU G UUGAGGUU 1369 AACCUCAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACCUCUUC 4830
86 GAGGUUAU G UCAAGCAU 1370 AUGCUUGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUAACCUC 4831
155 AAGAUAUU G UUAUCAUϋ 1371 AAUGAUAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUAUCUU 4832
221 AAAGACCU G UGAUAAAC 1372 GUUUAUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGUCUUU 4833
253 GGAAACGU G UGUCUAUA 1373 UAUAGACA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACGUUUCC 4834
255 AAACGUGU G UCUAUAUU 1374 AAUAUAGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACACGUUU 4835
273 UCAUAUCU G UAUAUAUA 1375 UAUAUAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAUAUGA 4836
344 AGGGAGAU G UACAGCAA 1376 UUGCUGUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCUCCCU 4837
373 AGAGUUCU G UGUUCAUC 1377 GAUGAACA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAACUCU 4838
375 AGUUCUGU G UUCAUCUϋ 1378 AAGAUGAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAGAACU 4839
457 AAGGCAUU G UCGUUGCA 1379 UGCAACGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUGCCUU 4840
478 ACCCCAAU G UGCCAGAA 1380 UUCUGGCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUGGGGU 4841
537 GCAUCUCU G UAUCUGUU 1381 AACAGAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAGAUGC 4842
543 CUGUAUCU G UUUGAAGC 1382 GCUUCAAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAUACAG 4843
580 UCAAAAAU G UUGCCAUU 1383 AAUGGCAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUUUGA 4844
625 CUGACUAU G UGAGACCA 1384 UGGUCUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUAGUCAG 4845
661 AUGCUGAU G UUCUGGUU 1385 AACCAGAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCAGCAU 4846
725 GGGCAACU G UGGAGAGA 1386 UCUCUCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUUGCCC 4847
814 AGGCAUUU G UCCAUGAG 1387 CUCAUGGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAUGCCU 4848
911 AGUAAGAU ,G UUCAGCAG 1388 CUGCUGAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCUUACU 4849
937 GUACAAAU G UAGUAAAG 1389 CUUUACUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUGUAC 4850
950 AAAGAAGU G UCAGGGAG 1390 CUCCCUGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUUCUUU 4851
965 AGGCAGCU G UUACACCA 1391 UGGUGUAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCUGCCU 4852
1019 AAAAGGAU G UGAGUUUG 1392 CAAACUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCCUUUU 4853
1027 GUGAGUUU G UUCUCCAA 1393 UUGGAGAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAACUCAC 4854
1065 UCUAUAAU G UUUGCACA 1394 UGUGCAAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUAUAGA 4855
1078 CACAACAU G UUGAUUCU 1395 AGAAUCAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUGUUGUG 4856
1100 UGAAUUCU G UACAGAAC 1396 GUUCUGUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAAUUCA 4857
1270 AAAGAAUϋ G UGUGUUUA 1397 UAAACACA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUUCUUU 4858
1272 AGAAUUGU G UGUUUAGU 1398 ACUAAACA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAAUUCU 4859
1274 AAUUGUGU G UUUAGUCC 1399 GGACUAAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACACAAUU 4860
1414 CUGCCCAU G UACAAAGU 1400 ACUUUGUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUGGGCAG 4861
1534 CAUUUACU G UGAUUAGG 1401 CCUAAUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUAAAUG 4862
1573 CUGAAAUU G UGCUGCUG 1402 CAGCAGCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUUUCAG 4863
1695 GAGGAGCU G UCCAAAAU 1403 AUUUUGGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCUCCUC 4864
1795 AUGGAGCU G UCUCUCAG 1404 CUGAGAGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCUCCAU 4865
1902 GACACUUU G UUUCUUAU 1405 AUAAGAAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAGUGUC 4866
1978 GUGGCUUU G UAGUGGAC 1406 GUCCACUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAGCCAC 4867
2086 CCCUGACU G UCACGUCC 1407 GGACGUGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUCAGGG 4868
2227 GGGCCAGU G UCACAGCC 1408 GGCUGUGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUGGCCC 4869
2320 AUGACGGU G UCUACUCA 1409 UGAGUAGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACCGUCAU 4870
2368 GAUACAGU G UAAAAGUG 1410 CACUUUUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUGUAUC 4871
2439 GGAGCACU G UACAUACC 1411 GGUAUGUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUGCUCC 4872
2512 AGGAUGAU G UUCAACAC 1412 GUGUUGAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCAUCCU 4873
2529 AAGCAAGU G UGUUUCAG 1413 CUGAAACA GGA GCCGUUAGGC UCCCUUCAAGGA' GCCGUUAGGC UCCGGG ACUUGCUU 4874
2531 GCAAGUGU G UUUCAGCA 1414 UGCUGAAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACACUUGC 4875
2563 GCUCAUUU G UGGCUUCU 1415 AGAAGCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAUGAGC 4876
2575 CUUCUGAU G UCCCAAAU 1416 AUUUGGGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCAGAAG 4877
2829 GUCUUUUU G UUUAAACC 1417 GGUUUAAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAAAGAC 4878
2890 UUCAGGCU G UUGAUAAG 1418 CUUAUCAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCCUGAA 4879
2943 GUAUCUUU G UUUAUUCC 1419 GGAAUAAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAGAUAC 4880
3002 UGCUCCUU G UCCUAAUA 1420 UAUUAGGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAGGAGCA 4881
3057 AAAAUUAU G UGGAAGUG 1421 CACUUCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUAAUUUU 4882
3084 CUGCAGCU G UCAAUAGC 1422 GCUAUUGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCUGCAG 4883
3109 GAAUUUUϋ G UCAGAUAA 1423 UUAUCUGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAAAUUC 4884
3166 UCUAAAAU G UAUUUUAG 1424 CUAAAAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUUAGA 4885
3182 GACUUCCU G UAGGGGGC 1425 GCCCCCUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGAAGUC 4886
3272 GACUUCCU G UAGGGGGC 1425 GCCCCCUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGAAGUC 4886
3203 UACUAAAU G UAUAUAGU 1426 ACUAUAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUAGUA 4887
3227 UACUAAAU G UAUUCCUG 1427 CAGGAAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUAGUA 4888
3235 GUAUUCCU G UAGGGGGC 1428 GCCCCCUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGAAUAC 4889
3256 UACUAAAU G UAUUUUAG 1429 CUAAAAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUAGUA 4890
15 UGCUUUUG G UACAAAUG 1430 CAUUUGUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAAAAGCA 4891
63 UAAGGGGA G CAUGAAGA 1431 UCUUCAUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCCCUUA 4892
73 AUGAAGAG G UGUUGAGG 1432 CCUCAACA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCUUCAU 4893
81 GUGUUGAG G UUAUGUCA 1433 UGACAUAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCAACAC 4894
91 UAUGUCAA G CAUCUGGC 1434 GCCAGAUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGACAUA 4895
98 AGCAUCUG G CACAGCUG 1435 CAGCUGUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGAUGCU 4896
103 CUGGCACA G CUGAAGGC 1436 GCCUUCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUGCCAG 4897
110 AGCUGAAG G CAGAUGGA 1437 UCCAUCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUCAGCU 4898
130 AUUUACAA G UACGCAAU 1438 AUUGCGUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGUAAAU 4899
182 AGACAAGA G CAAUAGUA 1439 UACUAUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUUGUCU 4900
188 GAGCAAUA G UAAAACAC 1440 GUGUUUUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAUUGCUC 4901
202 CACAUCAG G UCAGGGGG 1441 CCCCCUGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGAUGUG 4902
210 GUCAGGGG G UUAAAGAC 1442 GUCUUUAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCCUGAC 4903
242 UCCGAUAA G UUGGAAAC 1443 GUUUCCAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUAUCGGA 4904
251 UUGGAAAC G UGUGUCUA 1444 UAGACACA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUUUCCAA 4905
287 AUAUAAUG G UAAAGAAA 1445 UUϋCϋϋUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUAUAU 4906
305 ACACCUUC G UAACCCGC 1446 GCGGGUUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GAAGGUGU 4907
349 GAUGUACA G CAAUGGGG 1447 CCCCAUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUACAUC 4908
357 GCAAUGGG G CCAUUUAA 1448 UUAAAUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCAUUGC 4909
368 AUUUAAGA G UUCUGUGU 1449 ACACAGAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUUAAAU 4910
406 UAGAAGGG G CCCUGAGU 1450 ACUCAGGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCUUCUA 4911
413 GGCCCUGA G UAAUUCAC 1451 GUGAAUUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAGGGCC 4912
429 CUCAUUCA G CUGAACAA 1452 UUGUUCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAAUGAG 4913
443 CAACAAUG G CUAUGAAG 1453 CUUCAUAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUGUUG 4914
452 CUAUGAAG G CAUUGUCG 1454 CGACAAUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUCAUAG 4915
460 GCAUUGUC G UUGCAAUC 1455 GAUUGCAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GACAAUGC 4916
520 AGGACAUG G UGACCCAG 1456 CUGGGUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUGUCCU 4917
529 UGACCCAG G CAUCUCUG 1457 CAGAGAUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGGGUCA 4918
550 UGUUUGAA G CUACAGGA 1458 UCCUGUAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCAAACA 4919
561 ACAGGAAA G CGAUUUUA 1459 UAAAAUCG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUCCUGU 4920
616 AGACAAAG G CUGACUAU 1460 AUAGUCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUUGUCU 4921
667 AUGUUCUG G UUGCUGAG 1461 CUCAGCAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGAACAU 4922
675 GUUGCUGA G UCUACUCC 1462 GGAGUAGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAGCAAC 4923
689 UCCUCCAG G UAAUGAUG 1463 CAUCAUUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGGAGGA 4924
711 UACACUGA G CAGAUGGG 1464 CCCAUCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAGUGUA 4925
719 GCAGAUGG G CAACUGUG 1465 CACAGUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAUCUGC 4926
737 AGAGAAGG G UGAAAGGA 1466 UCCUUUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUUCUCU 4927
780 GGAAAAAA G UUAGCUGA 1467 UCAGCUAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUUUUCC 4928
784 AAAAGUUA G CUGAAUAU 1468 AUAUUCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAACUUUU 4929
803 ACCACAAG G UAAGGCAU 1469 AUGCCUUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUGUGGU 4930
808 AAGGUAAG G CAUUUGUC 1470 GACAAAUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUACCUU 4931
822 GUCCAUGA G UGGGCUCA 1471 UGAGCCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAUGGAC 4932
826 AUGAGUGG G CUCAUCUA 1472 UAGAUGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCACUCAU 4933
844 GAUGGGGA G UAUUUGAC 1473 GUCAAAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCCCAUC 4934
855 UUUGACGA G UACAAUAA 1474 UUAUUGUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCGUCAAA 4935
901 GAAUACAA G CAGUAAGA 1475 UCUUACUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGUAUUC 4936
904 UACAAGCA G UAAGAUGU 1476 ACAUCUUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCUUGUA 4937
916 GAUGUUCA G CAGGUAUU 1477 AAUACCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAACAUC 4938
920 UUCAGCAG G UAUUACUG 1478 CAGUAAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGCUGAA 4939
929 UAUUACUG G UACAAAUG 1479 CAUUUGUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGUAAUA 4940
940 CAAAUGUA G UAAAGAAG 1480 CUUCUUUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UACAUUUG 4941
948 GUAAAGAA G UGUCAGGG 1481 CCCUGACA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCUUUAC 4942
959 UCAGGGAG G CAGCUGUU 1482 AACAGCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCCCUGA 4943
962 GGGAGGCA G CUGUUACA 1483 UGUAACAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCCUCCC 4944
994 UCAAUAAA G UUACAGGA 1484 UCCUGUAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUAUUGA 4945
1023 GGAUGUGA G UUUGUUCU 1485 AGAACAAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCACAUCC 4946
1054 CGGAGAAG G CUUCUAUA 1486 UAUAGAAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUCUCCG 4947
1090 AUUCUAUA G UUGAAUUC 1487 GAAUUCAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAUAGAAU 4948
1126 ACAAAGAA G CUCCAAAC 1488 GUUUGGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCUUUGU 4949
1137 CCAAACAA G CAAAAUCA 1489 UGAUUUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGUUUGG 4950
1163 UCUCCGAA G CACAUGGG 1490 CCCAUGUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCGGAGA 4951
1174 CAUGGGAA G UGAUCCGU 1491 ACGGAUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCCCAUG 4952
1181 AGUGAUCC G UGAUUCUG 1492 CAGAAUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GGAUCACU 4953
1224 ACAACACA G CCACCAAA 1493 UUUGGUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUGUUGU 4954
1279 UGUGUUUA G UCCUUGAC 1494 GUCAAGGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAAACACA 4955
1298 AUCUGGAA G CAUGGCGA 1495 UCGCCAUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCCAGAU 4956
1303 GAAGCAUG G CGACUGGU 1496 ACCAGUCG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUGCUUC 4957
1310 GGCGACUG G UAACCGCC 1497 GGCGGUUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGUCGCC 4958
1336 UGAAUCAA G CAGGCCAG 1498 CUGGCCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGAUUCA 4959
1340 UCAAGCAG G CCAGCUUU 1499 AAAGCUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGCUUGA 4960
1344 GCAGGCCA G CUUUUCCU 1500 AGGAAAAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGCCUGC 4961
1363 UGCAGACA G UUGAGCUG 1501 CAGCUCAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUCUGCA 4962
1368 ACAGUUGA G CUGGGGUC 1502 GACCCCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAACUGϋ 4963
1374 GAGCUGGG G UCCUGGGU 1503 ACCCAGGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCAGCUC 4964
1381 GGUCCUGG G UUGGGAUG 1504 CAUCCCAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAGGACC 4965
1390 UUGGGAUG G UGACAUUU 1505 AAAUGUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUCCCAA 4966
1403 AUUUGACA G UGCUGCCC 1506 GGGCAGCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUCAAAU 4967
1421 UGUACAAA G UGAACUCA 1507 UGAGUUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUGUACA 4968
1442 GAUAAACA G UGGCAGUG 1508 CACUGCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUUUAUC 4969
1445 AAACAGUG G CAGUGACA 1509 UGUCACUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACUGUUU 4970
1448 CAGUGGCA G UGACAGGG 1510 CCCUGUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCCACUG 4971
1483 UACCUGCA G CAGCUUCA 1511 UGAAGCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCAGGUA 4972
1486 CUGCAGCA G CUUCAGGA 1512 UCCUGAAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCUGCAG 4973
1500 GGAGGGAC G UCCAUCUG 1513 CAGAUGGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUCCCUCC 4974
1511 CAUCUGCA G CGGGCUUC 1514 GAAGCCCG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCAGAUG 4975
1515 UGCAGCGG G CUUCGAUC 1515 GAUCGAAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCGCUGCA 4976
1525 UUCGAUCG G CAUUUACU 1516 AGUAAAUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CGAUCGAA 4977
1607 CACUAUAA G UGGGUGCU 1517 AGCACCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUAUAGUG 4978
1611 AUAAGUGG G UGCUUUAA 1518 UUAAAGCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCACUUAU 4979
1624 UUAACGAG G UCAAACAA 1519 UUGUUUGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCGUUAA 4980
1634 CAAACAAA G UGGUGCCA 1520 UGGCACCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUGUUUG 4981
1637 ACAAAGUG G UGCCAUCA 1521 UGAUGGCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACUUUGU 4982
1654 UCCACACA G UCGCUUUG 1522 CAAAGCGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUGUGGA 4983
1665 GCUUUGGG G CCCUCUGC 1523 GCAGAGGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCAAAGC 4984
1675 CCUCUGCA G CUCAAGAA 1524 UUCUUGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCAGAGG 4985
1692 CUAGAGGA G CUGUCCAA 1525 UUGGACAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCUCUAG 4986
1712 GACAGGAG G UUUACAGA 1526 UCUGUAAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCCUGUC 4987
1738 CAGAUCAA G UUCAGAAC 1527 GUUCUGAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGAUCUG 4988
1751 GAACAAUG G CCUCAUUG 1528 CAAUGAGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUGUUC 4989
1771 CUUUUGGG G CCCUUUCA 1529 UGAAAGGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCAAAAG 4990
1792 GAAAUGGA G CUGUCUCU 1530 AGAGACAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCAUUUC 4991
1803 GUCUCUCA G CGCUCCAU 1531 AUGGAGCG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAGAGAC 4992
1815 UCCAUCCA G CUUGAGAG 1532 CUCUCAAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGAUGGA 4993
1823 GCUUGAGA G UAAGGGAU 1533 AUCCCUUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUCAAGC 4994
1847 CCAGAACA G CCAGUGGA 1534 UCCACUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUUCUGG 4995
1851 AACAGCCA G UGGAUGAA 1535 UUCAUCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGCUGUU 4996
1862 GAUGAAUG G CACAGUGA 1536 UCACUGUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUCAUC 4997
1867 AUGGCACA G, UGAUCGUG 1537 CACGAUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUGCCAU 4998
1873 CAGUGAUC G UGGACAGC 1538 GCUGUCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GAUCACUG 4999
1880 , CGUGGACA G CACCGUGG 1539 CCACGGUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUCCACG 5000
1885 ACAGCACC G UGGGAAAG 1540 CUUUCCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GGUGCUGU 5001
1926 ACAACGCA G CCUCCCCA 1541 UGGGGAGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCGUUGU 5002
1.955 GGAUCCCA G UGGACAGA 1542 UCUGUCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGGAUCC 5003
1-965 .- GGACAGAA G CAAGGUGG 1543 CCACCUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCUGUCC 5004
1970 ' ' GAAGCAAG G UGGCUUUG 1544 CAAAGCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUGCUUC 5005
1973 GCAAGGUG G CUUUGUAG 1545 CUACAAAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACCUUGC 5006
1981 GCUUUGUA G UGGACAAA 1546 UUUGUCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UACAAAGC 5007
2002 CCAAAAUG G CCUACCUC 1547 GAGGUAGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUUUGG 5008
2021 AAUCCCAG G CAUUGCUA 1548 UAGCAAUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGGGAUU 5009
2032 UUGCUAAG G UUGGCACU 1549 AGUGCCAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUAGCAA 5010
2036 UAAGGUUG G CACUUGGA 1550 UCCAAGUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAACCUUA 5011
2051 GAAAUACA G UCUGCAAG 1551 CUUGCAGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUAUUUC 5012
2059 GUCUGCAA G CAAGCUCA 1552 UGAGCUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGCAGAC 5013
2063 GCAAGCAA G CUCACAAA 1553 UUUGUGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGCUUGC 5014
2091 ACUGUCAC G UCCCGUGC 1554 GCACGGGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUGACAGU 5015
2096 CACGUCCC G UGCGUCCA 1555 UGGACGCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GGGACGUG 5016
2100 UCCCGUGC G UCCAAUGC 1556 GCAUUGGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GCACGGGA 5017
2128 CAAUUACA G UGACUUCC 1557 GGAAGUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUAAUUG 5018
2156 GGACACCA G CAAAUUCC 1558 GGAAUUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGUGUCC 5019
2168 AUUCCCCA G CCCUCUGG 1559 CCAGAGGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGGGAAU 5020
2176 GCCCUCUG G UAGUUUAU 1560 AUAAACUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGAGGGC 5021
2179 CUCUGGUA G UUUAUGCA 1561 UGCAUAAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UACCAGAG 5022
2203 GCCAAGGA G CCUCCCCA 1562 UGGGGAGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCUUGGC 5023
2221 UUCUGAGG G CCAGUGUC 1563 GACACUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUGAGAA 5024
2225 CAGGGCCA G UGUCACAG 1564 CUGUGACA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGCCCUG 5025
2233 GUGUCACA G CCCUGAUU 1565 AAUCAGGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUGACAC 5026
2248 UUGAAUCA G UGAAUGGA 1566 UCCAUUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAUUCAA 5027
2263 GAAAAACA G UUACCUUG 1567 CAAGGUAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUUUUUC 5028
2290 AUAAUGGA G CAGGUGCU 1568 AGCACCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCAUUAU 5029
2294 UGGAGCAG G UGCUGAUG 1569 CAUCAGCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGCUCCA 5030
2318 GGAUGACG G UGUCUACU 1570 AGUAGACA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CGUCAUCC 5031
2331 UACUCAAG G UAUUUCAC 1571 GUGAAAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUGAGUA 5032
2357 CACGAAUG G UAGAUACA 1572 UGUAUCUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUCGUG 5033
2366 UAGAUACA G UGUAAAAG 1573 CUUUUACA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUAUCUA 5034
2374 GUGUAAAA G UGCGGGCU 1574 AGCCCGCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUUACAC 5035
2380 AAGUGCGG G CUCUGGGA 1575 UCCCAGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCGCACUU 5036
2392 UGGGAGGA G UUAACGCA 1576 UGCGUUAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCUCCCA 5037
2401 UUAACGCA G CCAGACGG 1577 CCGUCUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCGUUAA 5038
2413 GACGGAGA G UGAUACCC 1578 GGGUAUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUCCGUC 5039
2424 AUACCCCA G CAGAGUGG 1579 CCACUCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGGGUAU 5040
2429 CCAGCAGA G UGGAGCAC 1580 GUGCUCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUGCUGG 5041
2434 AGAGUGGA G CACUGUAC 1581 GUACAGUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCACUCU 5042
2450 CAUACCUG G CUGGAUUG 1582 CAAUCCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGGUAUG 5043
2523 CAACACAA G CAAGUGUG 1583 CACACUUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGUGUUG 5044
2527 ACAAGCAA G UGUGUUUC 1584 GAAACACA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGCUUGU 5045
2537 GUGUUUCA G CAGAACAU 1585 AUGUUCUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAAACAC 5046
2555 CUCGGGAG G CUCAUUUG 1586 CAAAUGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCCCGAG 5047
2566 CAUUUGUG G CUUCUGAU 1587 AUCAGAAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACAAAUG 5048
2612 CCCACCUG G CCAAAUCA 1588 UGAUUUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGGUGGG 5049
2632 ACCUGAAG G CGGAAAUU 1589 AAUUUCCG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUCAGGU 5050
2648 UCACGGGG G CAGUCUCA 1590 UGAGACUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCCGUGA 5051
2651 CGGGGGCA G UCUCAUUA 1591 UAAUGAGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCCCCCG 5052
2674 CUUGGACA G CUCCUGGG 1592 CCCAGGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUCCAAG 5053
2704 AUGGAACA G CUCACAAG 1593 CUUGUGAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUUCCAU 5054
2712 GCUCACAA G UAUAUCAU 1594 AUGAUAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGUGAGC 5055
2729 UCGAAUAA G UACAAGUA 1595 UACUUGUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUAUUCGA 5056
2735 AAGUACAA G UAUUCUUG 1596 CAAGAAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGUACUU 5057
2757 AGAGACAA G UUCAAUGA 1597 UCAUUGAA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGUCUCU 5058
2776 CUCUUCAA G UGAAUACU 1598 AGUAUUCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGAAGAG 5059
2806 CAAAGGAA G CCAACUCU 1599 AGAGUUGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCCUUUG 5060
2821 CUGAGGAA G UCUUUUUG 1600 CAAAAAGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCCUCAG 5061
2861 UGAAAAUG G CACAGAUC 1601 GAUCUGUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUUUCA 5062
2887 CUAUUCAG G CUGUUGAU 1602 AUCAACAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGAAUAG 5063
2899 UUGAUAAG G UCGAUCUG 1603 CAGAUCGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUAUCAA 5064
2935 UUGCACGA G UAUCUUUG 1604 CAAAGAUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCGUGCAA 5065
2978 GACACCUA G UCCUGAUG 1605 CAUCAGGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAGGUGUC 5066
2991 GAUGAAAC G UCUGCUCC 1606 GGAGCAGA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUUUCAUC 5067
3023 UAUCAACA G CACCAUUC 1607 GAAUGGUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUUGAUA 5068
3035 CAUUCCUG G CAUUCACA 1608 UGUGAAUG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGGAAUG 5069
3063 AUGUGGAA G UGGAUAGG 1609 CCUAUCCA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCCACAU 5070
3081 GAACUGCA G CUGUCAAU 1610 AUUGACAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCAGUUC 5071
3091 UGUCAAUA G CCUAGGGC 1611 GCCCUAGG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAUUGACA 5072
3098 AGCCUAGG G CUGAAUUU 1612 AAAUUCAG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUAGGCU 5073
3189 UGUAGGGG G CGAUAUAC 1613 GUAUAUCG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCCUACA 5074
3242 UGUAGGGG G CGAUAUAC 1613 GUAUAUCG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCCUACA 5074
3210 UGUAUAUA G UACAUUUA 1614 UAAAUGUA GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAUAUACA 5075
3279 UGUAGGGG G CGAUAAAA 1615 UUUUAUCG GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCCUACA 5076
14 AUGCUUUU G GUACAAAU 1868 AUUUGUAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAAGGAU 5077
23 GUACAAAU G GAUGUGGA 1869 UCCACAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUGUAC 5078
24 UACAAAUG G AUGUGGAA 1870 UUCCACAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUUGUA 5079
29 AUGGAUGU G GAAUAUAA 1871 UUAUAUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAUCCAU 5080
30 UGGAUGUG G AAUAUAAU 1872 AUUAUAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACAUCCA 5081
58 UUGUUUAA G GGGAGCAU 1873 AUGCUCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUAAACAA 5082
59 UGUUUAAG G GGAGCAUG 1874 CAUGCUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUAAACA 5083
60 GUUUAAGG G GAGCAUGA 1875 UCAUGCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUUAAAC 5084
61 UUUAAGGG G AGCAUGAA 1876 UUCAUGCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCUUAAA 5085
70 AGCAUGAA G AGGUGUUG 1877 CAACACCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCAUGCU 5086
72 CAUGAAGA G GUGUUGAG 1878 CUCAACAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUUCAUG 5087
80 GGUGUUGA G GUUAUGUC 1879 FGACAUAAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAACACC 5088
97 AAGCAUCU G GCACAGCU 1880 AGCUGUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAUGCUU 5089
109 CAGCUGAA G GCAGAUGG 1881 CCAUCUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCAGCUG 5090
113 UGAAGGCA G AUGGAAAU 1882 AUUUCCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCCUUCA 5091
116 AGGCAGAU G GAAAUAUU 1883 AAUAUUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCUGCCU 5092
117 GGCAGAUG G AAAUAUUU 1884 AAAUAUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUCUGCC 5093
143 CAAUUUGA G ACUAAGAU 1885 AUCUUAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAAAUUG 5094
149 GAGACUAA G AUAUUGUU 1886 AACAAUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUAGUCUC 5095
175 CUAUUGAA G ACAAGAGC 1887 GCUCUUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCAAUAG 5096
180 GAAGACAA G AGCAAUAG 1888 CUAUUGCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGUCUUC 5097
201 ACACAUCA G GUCAGGGG 1889 CCCCUGAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAUGUGU 5098
206 UCAGGUCA G GGGGUUAA 1890 UUAACCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC ΌCCGGG UGACCUGA 5099
207 CAGGUCAG G GGGUUAAA 1891 UUUAACCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGACCUG 5100
208 AGGUCAGG G GGUUAAAG 1892 CUUUAACC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUGACCU 5101
209 GGUCAGGG G GUUAAAGA 1893 UCUUUAAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCUGACC 5102
216 GGGUUAAA G ACCUGUGA 1894 UCACAGGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUAACCC 5103
245 GAUAAGUU G GAAACGUG 1895 CACGUUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AACUUAUC 5104
246 AUAAGUUG G AAACGUGU 1896 ACACGUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAACUUAU 5105
286 UAUAUAAU G GUAAAGAA 1897 UUCUUUAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUAUAUA 5106
292 AUGGUAAA G AAAGACAC 1898 GUGUCUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUACCAU 5107
296 UAAAGAAA G ACACCUUC 1899 GAAGGUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUCUUUA 5108
324 UUUCCAAA G AGAGGAAU 1900 AUUCCUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUGGAAA 5109
326 UCCAAAGA G AGGAAUCA 1901 UGAUUCCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUUUGGA 5110
328 CAAAGAGA G GAAUCACA 1902 UGUGAUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUCUUUG 5111
329 AAAGAGAG G AAUCACAG 1903 CUGUGAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCUCUUU 5112
337 GAAUCACA G GGAGAUGU 1904 ACAUCUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUGAUUC 5113
338 AAUCACAG G GAGAUGUA 1905 UACAUCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGUGAUU 5114
339 AUCACAGG G AGAUGUAC 1906 GUACAUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUGUGAU 5115
341 CACAGGGA G AUGUACAG 1907 CUGUACAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCCUGUG 5116
354 ACAGCAAU G GGGCCAUU 1908 AAUGGCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUGCUGU 5117
355 CAGCAAUG G GGCCAUUU 1909 AAAUGGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUGCUG 5118
356 AGCAAUGG G GCCAUUUA 1910 UAAAUGGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAUUGCU 5119
366 CCAUUUAA G AGUUCUGU 1911 ACAGAACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUAAAUGG 5120
400 ACCUUCUA G AAGGGGCC 1912 GGCCCCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAGAAGGU 5121
403 UUCUAGAA G GGGCCCUG 1913 CAGGGCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCUAGAA 5122
404 UCUAGAAG G GGCCCUGA 1914 UCAGGGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUCUAGA 5123
405 CUAGAAGG G GCCCUGAG 1915 CUCAGGGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUUCUAG 5124
442 ACAACAAU G GCUAUGAA 1916 UUCAUAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUGUUGU 5125
451 GCUAUGAA G GCAUUGUC 1917 GACAAUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCAUAGC 5125
484 AUGUGCCA G AAGAUGAA 1918 UUCAUCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGGACAU 5127
487 UGCCAGAA G AUGAAACA 1919 UGUUUCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCUGGCA 5128
513 CAAAUAAA G GACAUGGU 1920 ACCAUGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUAUUUG 5129
514 AAAUAAAG G ACAUGGUG 1921 CACCAUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUUAUUU 5130
519 AAGGACAU G GUGACCCA 1922 UGGGUCAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUGUCCUU 5131
528 GUGACCCA G GCAUCUCU 1923 AGAGAUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGGUCAC 5132
556 AAGCUACA G GAAAGCGA 1924 UCGCUUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUAGCUU 5133
557 AGCUACAG G AAAGCGAU 1925 AUCGCUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGUAGCU 5134
605 UGAAACAU G GAAGACAA 1926 UUGUCUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUGUUUCA 5135
606 GAAACAUG G AAGACAAA 1927 UUUGUCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUGUUUC 5136
609 ACAUGGAA G ACAAAGGC 1928 GCCUUUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCCAUGU 5137
615 AAGACAAA G GCUGACUA 1929 ■ UAGUCAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUGUCUU 5138
629 CUAUGUGA G ACCAAAAC 1930 GUUUUGGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCACAUAG 5139
642 AAACUUGA G ACCUACAA 1931 UUGUAGGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAAGUUU 5140
666 GAUGUUCU G GUUGCUGA 1932 UCAGCAAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAACAUC 5141
688 CUCCUCCA G GUAAUGAU 1933 AUCAUUAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGAGGAG 5142
714 ACUGAGCA G AUGGGCAA 1934 UUGCCCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCUCAGU 5143
717 GAGCAGAU G GGCAACUG 1935 CAGUUGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCUGCUC 5144
718 AGCAGAUG G GCAACUGU 1936 ACAGUUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUCUGCU 5145
727 GCAACUGU G GAGAGAAG 1937 CUUCUCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAGUUGC 5146
728 CAACUGUG G AGAGAAGG 1938 CCUUCUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACAGUUG 5147
730 ACUGUGGA G AGAAGGGU 1939 ACCCUUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCACAGU 5148
732 UGUGGAGA G AAGGGUGA 1940 UCACCCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUCCACA 5149
735 GGAGAGAA G GGUGAAAG 1941 CUUUCACC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCUCUCC 5150
736 GAGAGAAG G GUGAAAGG 1942 CCUUUCAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUCUCUC 5151
743 GGGUGAAA G GAUCCACC 1943 GGUGGAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUCACCC 5152
744 GGUGAAAG G AUCCACCU 1944 AGGUGGAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUUCACC 5153
772 UCAUUGCA G GAAAAAAG 1945 CUUUUUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCAAUGA 5154
773 CAUUGCAG G AAAAAAGU 1946 ACUUUϋUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGCAAUG 5155
793 CUGAAUAU G GACCACAA 1947 ' UUGUGGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUAUUCAG 5156
794 UGAAUAUG G ACCACAAG 1948 CUUGUGGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUAUUCA 5157
802 GACCACAA G GUAAGGCA 1949 UGCCUUAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGUGGUC 5158
807 CAAGGUAA G GCAUUUGU 1950 ACAAAUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUACCUUG 5159
824 CCAUGAGU G GGCUCAUC 1951 GAUGAGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUCAUGG 5160
825 CAUGAGUG G GCUCAUCU 1952 AGAUGAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACUCAUG 5161
839 UCUACGAU G GGGAGUAU 1953 AUACUCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCGUAGA 5162
840 CUACGAUG G GGAGUAUU 1954 AAUACUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUCGUAG 5163
841 UACGAUGG G GAGUAUUU 1955 AAAUACUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAUCGUA 5164
842 ACGAUGGG G AGUAUUUG 1956 CAAAUACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCAUCGU 5165
870 AAUGAUGA G AAAUUCUA 1957 UAGAAUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAUCAUU 5166
889 UAUCCAAU G GAAGAAUA 1958 UAUUCUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUGGAUA 5167
890 AUCCAAUG G AAGAAUAC 1959 GUAUUCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUGGAU 5168
893 CAAUGGAA G AAUACAAG 1960 CUUGUAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCCAUUG 5169
908 AGCAGUAA G AUGUUCAG 1961 CUGAACAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUACUGCU 5170
919 GUUCAGCA G GUAUUACU 1962 AGUAAUAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCUGAAC 5171
928 GUAUUACU G GUACAAAU 1963 AUUUGUAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUAAUAC 5172
945 GUAGUAAA G AAGUGUCA 1964 UGACACUϋ GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUACUAC 5173
954 AAGUGUCA G GGAGGCAG 1965 CUGCCUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGACACUU 5174
955 AGUGUCAG G GAGGCAGC 1966 GCUGCCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGACACU 5175
956 GUGUCAGG G AGGCAGCU 1967 AGCUGCCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUGACAC 5176
958 GUCAGGGA G GCAGCUGU 1968 ACAGCUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCCUGAC 5177
977 CACCAAAA G AUGCACAU 1969 AUGUGCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUUGGUG 5178
1000 AAGUUACA G GACUCUAU 1970 AUAGAGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUAACUU 5179
1001 AGUUACAG G ACUCUAUG 1971 CAUAGAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGUAACU 5180
1015 AUGAAAAA G GAUGUGAG 1972 CUCACAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUUUCAU 5181
1016 UGAAAAAG G AUGUGAGU 1973 ACUCACAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUUUUCA 5182
1044 UCCCGCCA G ACGGAGAA 1974 UUCUCCGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGCGGGA 5183
1047 CGCCAGAC G GAGAAGGC 1975 GCCUUCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUCUGGCG 5184
1048 GCCAGACG G AGAAGGCU 1976 AGCCUUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CGUCUGGC 5185
1050 CAGACGGA G AAGGCUUC 1977 GAAGCCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCGUCUG 5186
1053 ACGGAGAA G GCUUCUAU 1978 AUAGAAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCUCCGU 5187
1105 UCUGUACA G AACAAAAC 1979 GUUUUGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUACAGA 5188
1123 ACAACAAA G AAGCUCCA 1980 UGGAGCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUGUUGU 5189
1169 AAGCACAU G GGAAGUGA 1981 UCACUUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUGUGCUU 5190
1170 AGCACAUG G GAAGUGAU 1982 AUCACUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUGUGCU 5191
1171 GCACAUGG G AAGUGAUC 1983 GAUCACUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAUGUGC 5192
1191 GAUUCUGA G GACUUUAA 1984 UUAAAGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAGAAUC 5193
1192 AUUCUGAG G ACUUUAAG 1985 CUUAAAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCAGAAU 5194
1200 GACUUUAA G AAAACCAC 1986 GUGGUUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUAAAGUC 5195
1254 UUGCUGCA G AUUGGACA 1987 UGUCCAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCAGCAA 5196
1258 UGCAGAUU G GACAAAGA 1988 UCUUUGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAUCUGCA 5197
1259 GCAGAUUG G ACAAAGAA 1989 UUCUUUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAAUCUGC 5198
1265 UGGACAAA G AAUUGUGU 1990 ACACAAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUGUCCA 5199
1294 ACAAAUCU G GAAGCAUG 1991 CAUGCUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAUUUGU 5200
1295 CAAAUCUG G AAGCAUGG 1992 CCAUGCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGAUUUG 5201
1302 GGAAGCAU G GCGACUGG 1993 CCAGUCGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUGCUUCC 5202
1309 UGGCGACU G GUAACCGC 1994 GCGGUUAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUCGCCA 5203
1339 AUCAAGCA G GCCAGCUU 1995 AAGCUGGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCUUGAU 5204
1359 CUGCUGCA G ACAGUUGA 1996 UCAACUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCAGCAG 5205
1371 GUUGAGCU G GGGUCCUG 1997 CAGGACCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCUCAAC 5206
1372 UUGAGCUG G GGUCCUGG 1998 CCAGGACC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGCUCAA 5207
1373 UGAGCUGG G GUCCUGGG 1999 CCCAGGAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAGCUCA 5208
1379 GGGGUCCU G GGUUGGGA 2000 UCCCAACC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGACCCC 5209
1380 GGGUCCUG G GUUGGGAU 2001 AUCCCAAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGGACCC 5210
1384 CCUGGGUU G GGAUGGUG 2002 CACCAUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AACCCAGG 5211
1385 CUGGGUUG G GAUGGUGA 2003 UCACCAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAACCCAG 5212
1386 UGGGUUGG G AUGGUGAC 2004 GUCACCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAACCCA 5213
1389 GUUGGGAU G GUGACAUU 2005 AAUGUCAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCCCAAC 5214
1434 CUCAUACA G AUAAACAG 2006 CUGUUUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUAUGAG 5215
1444 UAAACAGU G GCAGUGAC 2007 GUCACUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUGUUUA 5216
1454 CAGUGACA G GGACACAC 2008 GUGUGUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUCACUG 5217
1455 AGUGACAG G GACACACU 2009 AGUGUGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGUCACU 5218
1456 GUGACAGG G ACACACUC 2010 GAGUGUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUGUCAC 5219
1472 CGCCAAAA G AUUACCUG 2011 CAGGUAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUUGGCG 5220
1492 CAGCUUCA G GAGGGACG 2012 CGUCCCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAAGCUG 5221
1493 AGCUUCAG G AGGGACGU 2013 ACGUCCCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGAAGCU 5222
1495 CUUCAGGA G GGACGUCC 2014 GGACGUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCUGAAG 5223
1496 UUCAGGAG G GACGUCCA 2015 UGGACGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCCUGAA 5224
1497 UCAGGAGG G ACGUCCAU 2016 AUGGACGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUCCUGA 5225
1513 UCUGCAGC G GGCUUCGA 2017 UCGAAGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GCUGCAGA 5226
1514 CUGCAGCG G GCUUCGAU 2018 AUCGAAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CGCUGCAG 5227
1524 CUUCGAUC G GCAUUUAC 2019 GUAAAUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GAUCGAAG 5228
1541 UGUGAUUA G GAAGAAAU 2020 AUUUCUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAAUCACA 5229
1542 GUGAUUAG G AAGAAAUA 2021 UAUUUCUϋ GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUAAUCAC 5230
1545 AUUAGGAA G AAAUAUCC 2022 GGAUAUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCCUAAU 5231
1561 CAACUGAU G GAUCUGAA 2023 UUCAGAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCAGUUG 5232
1562 AACUGAUG G AUCUGAAA 2024 UUUCAGAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUCAGUU 5233
1584 CUGCUGAC G GAUGGGGA 2025 UCCCCAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUCAGCAG 5234
1585 UGCUGACG G AUGGGGAA 2026 UUCCCCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CGUCAGCA 5235
1588 UGACGGAU G GGGAAGAC 2027 GUCUUCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUCCGUCA 5236
1589 GACGGAUG G GGAAGACA 2028 UGUCUUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUCCGUC 5237
1590 ACGGAUGG G GAAGACAA 2029 UUGUCUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAUCCGU 5238
1591 CGGAUGGG G AAGACAAC 2030 GUUGUCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCAΌCCG 5239
1594 AUGGGGAA G ACAACACU 2031 AGUGUUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCCCCAU 5240
1609 CUAUAAGU G GGUGCUUU 2032 AAAGCACC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUUAUAG 5241
1610 UAUAAGUG G GUGCUUUA 2033 UAAAGCAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACUUAUA 5242
1623 UUUAACGA G GUCAAACA 2034 UGUUUGAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCGUUAAA 5243
1636 AACAAAGU G GUGCCAUC 2035 GAUGGCAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUUUGUU 5244
1662 GUCGCUUU G GGGCCCUC 2036 GAGGGCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAGCGAC 5245
1663 UCGCUUUG G GGCCCUCU 2037 AGAGGGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAAAGCGA 5246
1664 CGCUUUGG G GCCCUCUG 2038 CAGAGGGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAAAGCG 5247
1681 CAGCUCAA G AACUAGAG 2039 CUCUAGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGAGCUG 5248
1687 AAGAACUA G AGGAGCUG 2040 CAGCUCCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAGUUCUU 5249
1689 GAACUAGA G GAGCUGUC 2041 GACAGCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUAGUUC 5250
1690 AACUAGAG G AGCUGUCC 2042 GGACAGCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCUAGUU 5251
1708 AAAUGACA G GAGGUUUA 2043 UAAACCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUCAUUU 5252
1709 AAUGACAG G AGGUUUAC 2044 GUAAACCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGUCAUU 5253
1711 UGACAGGA G GUUUACAG 2045 CUGUAAAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCUGUCA 5254
1719 GGUUUACA G ACAUAUGC 2046 GCAUAUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUAAACC 5255
1732 AUGCUUCA G AUCAAGUU 2047 AACUUGAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAAGCAU 5256
1743 CAAGUUCA G AACAAUGG 2048 CCAUUGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAACUUG 5257
1750 AGAACAAU G GCCUCAUU 2049 AAUGAGGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUGUUCU 5258
1768 AUGCUUUU G GGGCCCUU 2050 AAGGGCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAAAGCAU 5259
1769 UGCUUUUG G GGCCCUUU 2051 AAAGGGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAAAAGCA 5260
1770 GCUUUUGG G GCCCUUUC 2052 GAAAGGGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAAAAGC 5261
1783 UUUCAUCA G GAAAUGGA 2053 UCCAUUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAUGAAA 5262
1784 UUCAUCAG G AAAUGGAG 2054 CUCCAUUϋ GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGAUGAA 5263
1789 CAGGAAAU G GAGCUGUC 2055 GACAGCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUCCUG 5264
1790 AGGAAAUG G AGCUGUCU 2056 AGACAGCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUUCCU 5265
1821 CAGCUUGA G AGUAAGGG 2057 CCCUUACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAAGCUG 5266
1827 GAGAGUAA G GGAUUAAC 2058 GUUAAUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUACUCUC 5267
1828 AGAGUAAG G GAUUAACC 2059 GGUUAAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUACUCU 5268
1829 GAGUAAGG G AUUAACCC 2060 GGGUUAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUUACUC 5269
1842 ACCCUCCA G AACAGCCA 2061 UGGCUGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGAGGGU 5270
1853 CAGCCAGU G GAUGAAUG 2062 CAUUCAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUGGCUG 5271
1854 AGCCAGUG G AUGAAUGG 2063 CCAUUCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACUGGCU 5272
1861 GGAUGAAU G GCACAGUG 2064 CACUGUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUCAUCC 5273
1875 GUGAUCGU G GACAGCAC 2065 GUGCUGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACGAUCAC 5274
1876 UGAUCGUG G ACAGCACC 2066 GGUGCUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACGAUCA 5275
1887 AGCACCGU G GGAAAGGA 2067 UCCUUUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACGGUGCU 5276
1888 GCACCGUG G GAAAGGAC 2068 GUCCUUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACGGUGC 5277
1889 CACCGUGG G AAAGGACA 2069 UGUCCUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCACGGUG 5278
1893 GUGGGAAA G GACACUUϋ 2070 AAAGUGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUCCCAC 5279
1894 UGGGAAAG G ACACUUUG 2071 CAAAGUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUUCCCA 5280
1916 UAUCACCU G GACAACGC 2072 GCGUUGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGUGAUA 5281
1917 AUCACCUG G ACAACGCA 2073 UGCGUUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGGUGAU 5282
1946 CCUUCUCU G GGAUCCCA 2074 UGGGAUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAGAAGG 5283
1947 CUUCUCUG G GAUCCCAG 2075 CUGGGAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GAGAGAAG 5284
1948 UUCUCUGG G AUCCCAGU 2076 ACUGGGAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAGAGAA 5285
1957 AUCCCAGU G GACAGAAG 2077 CUUCUGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUGGGAU 5286
1958 UCCCAGUG G ACAGAAGC 2078 GCUUCUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACUGGGA 5287
1962 AGUGGACA G AAGCAAGG 2079 CCUUGCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUCCACU 5288
1969 AGAAGCAA G GUGGCUUU 2080 AAAGCCAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGCUUCU 5289
1972 AGCAAGGU G GCUUUGUA 2081 UACAAAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACCUUGCU 5290
1983 UUUGUAGU G GACAAAAA 2082 UUUUUGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUACAAA 5291
1984 UUGUAGUG G ACAAAAAC 2083 GUUUUUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACUACAA 5292
2001 ACCAAAAU G GCCUACCϋ 2084 AGGUAGGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUUGGU 5293
2020 AAAUCCCA G GCAUUGCU 2085 AGCAAUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGGAUUU 5294
2031 AUUGCUAA G GUUGGCAC 2086 GUGCCAAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUAGCAAU 5295
2035 CUAAGGUU G GCACUUGG 2087 CCAAGUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AACCUUAG 5296
2042 UGGCACUU G GAAAUACA 2088 UGUAUUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAGUGCCA 5297
2043 GGCACUUG G AAAUACAG 2089- CUGUAUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAAGUGCC 5298
2148 ACGAACAA G GACACCAG 2090 CUGGUGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGUUCGU 5299
2149 CGAACAAG G ACACCAGC 2091 GCUGGUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUGUUCG 5300
2175 AGCCCUCU G GUAGUUUA 2092 UAAACUAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAGGGCU 5301
2200 UUCGCCAA G GAGCCUCC _j 2093 GGAGGCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGGCGAA 5302
2201 UCGCCAAG G AGCCUCCC 2094 GGGAGGCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUGGCGA 5303
2219 AAUUCUCA G GGCCAGUG 2095 CACUGGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAGAAUU 5304
2220 AUUCUCAG G GCCAGUGU 2096 ACACUGGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUGAGAAU 5305
2254 CAGUGAAU G GAAAAACA 2097 UGUUUUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUCACUG 5306
2255 AGUGAAUG G AAAAACAG 2098 CUGUUUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUCACU 5307
2271 GUUACCUU G GAACUACU 2099 AGUAGUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAGGUAAC 5308
2272 UUACCUUG G AACUACUG 2100 CAGUAGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAAGGUAA 5309
2280 GAACUACU G GAUAAUGG 2101 CCAUUAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGUAGUUC 5310
2281 AACUACUG G AUAAUGGA 2102 UCCAUUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGUAGUU 5311
2287 UGGAUAAU G GAGCAGGU 2103 ACCUGCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUAUCCA 5312
2288 GGAUAAUG G AGCAGGUG 2104 CACCUGCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUAUCC 5313
2293 AUGGAGCA G GUGCUGAU 2105 AUCAGCAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCUCCAU 5314
2310 GCUACUAA G GAUGACGG 2106 CCGUCAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUAGUAGC 5315
2311 CUACUAAG G AUGACGGU 2107 ACCGUCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUAGUAG 5316
2317 AGGAUGAC G GUGUCUAC 2108 GUAGACAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUCAUCCU 5317
2330 CUACUCAA G GUAUUUCA 2109 UGAAAUAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGAGUAG 5318
2356 ACACGAAU G GUAGAUAC 2110 GUAUCUAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUCGUGU 5319
2360 GAAUGGUA G AUACAGUG 2111 CACUGUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UACCAUUC 5320
2378 AAAAGUGC G GGCUCUGG 2112 CCAGAGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GCACUUUU 5321
2379 AAAGUGCG G GCUCUGGG 2113 CCCAGAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CGCACUUU 5322
2385 CGGGCUCU G GGAGGAGU 2114 ACUCCUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGAGCCCG 5323
2386 GGGCUCUG G GAGGAGUU 2115 AACUCCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGAGCCC 5324
2387 GGCUCUGG G AGGAGUUA 2116 UAACUCCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAGAGCC 5325
2389 CUCUGGGA G GAGUUAAC 2117 GUUAACUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCCAGAG 5326
2390 UCUGGGAG G AGUUAACG 2118 CGUUAACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCCCAGA 5327
2405 CGCAGCCA G ACGGAGAG 2119 CUCUCCGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGCUGCG 5328
2408 AGCCAGAC G GAGAGUGA 2120 UCACUCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUCUGGCU 5329
2409 GCCAGACG G AGAGUGAU 2121 AUCACUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CGUCUGGC 5330
2411 CAGACGGA G AGUGAUAC 2122 GUAUCACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCGUCUG 5331
2427 CCCCAGCA G AGUGGAGC 2123 GCUCCACU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCUGGGG 5332
2431 AGCAGAGU G GAGCACUG 2124 CAGUGCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUCUGCU 5333
2432 GCAGAGUG G AGCACUGU 2125 ACAGUGCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACUCUGC 5334
2449 ACAUACCU G GCUGGAUU 2126 AAUCCAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGUAUGU 5335
2453 ACCUGGCU G GAUUGAGA 2127 UCUCAAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGCCAGGU 5336
2454 CCUGGCUG G AUUGAGAA 2128 UUCUCAAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGCCAGG 5337
2460 UGGAUUGA G AAUGAUGA 2129 UCAUCAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAAUCCA 5338
2477 AAUACAAU G GAAUCCAC 2130 GUGGAUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUGUAUU 5339
2478 AUACAAUG G AAUCCACC 2131 GGUGGAUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUUGUAU 5340
2489 UCCACCAA G ACCUGAAA 2132 UUUCAGGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUGGUGGA 5341
2505 AUUAAUAA G GAUGAUGU 2133 ACAUCAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUAUUAAU 5342
2506 UUAAUAAG G AUGAUGUU 2134 AACAUCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUAUUAA 5343
2540 UUUCAGCA G AACAUCCU 2135 AGGAUGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGCUGAAA 5344
2550 ACAUCCUC G GGAGGCUC 2136 GAGCCUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GAGGAUGU 5345
2551 CAUCCUCG G GAGGCUCA 2137 UGAGCCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CGAGGAUG 5346
2552 AUCCUCGG G AGGCUCAU 2138 AUGAGCCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCGAGGAU 5347
2554 CCUCGGGA G GCUCAUUU 2139 AAAUGAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCCGAGG 5348
2565 UCAUUUGU G GCUUCUGA 2140 UCAGAAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAAAUGA 5349
2611 UCCCACCU G GCCAAAUC 2141 GAUUUGGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGUGGGA 5350
2631 GACCUGAA G GCGGAAAU 2142 AUUUCCGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUCAGGUC 5351
2634 CUGAAGGC G GAAAUUCA 2143 UGAAUUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GCCUUCAG 5352
2635 UGAAGGCG G AAAUUCAC 2144 GUGAAUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CGCCUUCA 5353
2644 AAAUUCAC G GGGGCAGU 2145 ACUGCCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG GUGAAUUU 5354
2645 AAUUCACG G GGGCAGUC 2146 GACUGCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CGUGAAUU 5355
2646 AUUCACGG G GGCAGUCU 2147 AGACUGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCGUGAAU 5356
2647 UUCACGGG G GCAGUCUC 2148 GAGACUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCGUGAA 5357
2669 UCUGACUU G GACAGCUC 2149 GAGCUGUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AAGUCAGA 5358
2670 CUGACUUG G ACAGCUCC 2150 GGAGCUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAAGUCAG 5359
2680 CAGCUCCU G GGGAUGAU 2151 AUCAUCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGAGCUG 5360
2681 AGCUCCUG G GGAUGAUU 2152 AAUCAUCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAGGAGCU 5361
2682 GCUCCUGG G GAUGAUUA 2153 UAAUCAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCAGGAGC 5362
2683 CUCCUGGG G AUGAUUAU 2154 AUAAUCAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCAGGAG 5363
2698 AUGACCAU G GAACAGCU 2155 AGCUGUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUGGUCAU 5364
2699 UGACCAUG G AACAGCUC 2156 GAGCUGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CAUGGUCA 5365
2750 UGAUCUCA G AGACAAGU 2157 ACUUGUCϋ GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAGAUCA 5366
2752 AUCUCAGA G ACAAGUUC 2158 GAACUUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUGAGAU 5367
2802 AUCCCAAA G GAAGCCAA 2159 UUGGCUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUUGGGAU 5368
2803 UCCCAAAG G AAGCCAAC 2160 GUUGGCUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUUUGGGA 5369
2817 AACUCUGA G GAAGUCUϋ 2161 AAGACUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCAGAGUU 5370
2818 ACUCUGAG G AAGUCUUU 2162 AAAGACUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUCAGAGU 5371
2839 UUAAACCA G AAAACAUU 2163 AAUGUUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGUUUAA 5372
2860 UUGAAAAU G GCACAGAU 2164 AUCUGUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AUUUUCAA 5373
2866 AUGGCACA G AUCUUUUC 2165 GAAAAGAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUGCCAU 5374
2886 GCUAUUCA G GCUGUUGA 2166 UCAACAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAAUAGC 5375
2898 GUUGAUAA G GUCGAUCU 2167 AGAUCGAC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UUAUCAAC 5376
2914 UGAAAUCA G AAAUAUCC 2168 GGAUAUUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGAUUUCA 5377
2958 CCUCCACA G ACUCCGCC 2169 GGCGGAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGUGGAGG 5378
2968 CUCCGCCA G AGACACCU 2170 AGGUGUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGGCGGAG 5379
2970 CCGCCAGA G ACACCUAG 2171 CUAGGUGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCUGGCGG 5380
3034 CCAUUCCU G GCAUUCAC 2172 GUGAAUGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG AGGAAUGG 5381
3059 AAUUAUGU G GAAGUGGA 2173 UCCACUUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACAUAAUU 5382
3060 AUUAUGUG G AAGUGGAU 2174 AUCCACUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACAUAAU 5383
3065 GUGGAAGU G GAUAGGAG 2175 CUCCUAUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG ACUUCCAC 5384
3066 UGGAAGUG G AUAGGAGA 2176 UCUCCUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CACUUCCA 5385
3070 AGUGGAUA G GAGAACUG 2177 CAGUUCUC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAUCCACU 5386
3071 GUGGAUAG G AGAACUGC 2178 GCAGUUCU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUAUCCAC 5387
3073 GGAUAGGA G AACUGCAG 2179 CUGCAGUU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UCCUAUCC 5388
3096 AUAGCCUA G GGCUGAAU 2180 AUUCAGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAGGCUAU 5389
3097 UAGCCUAG G GCUGAAUU 2181 AAUUCAGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUAGGCUA 5390
3113 UUUUGUCA G AUAAAUAA 2182 UUAUUUAU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UGACAAAA 5391
3174 GUAUUUUA G ACUUCCUG 2183 CAGGAAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAAAAUAC 5392
3264 GUAUUUUA G ACUUCCUG 2183 CAGGAAGU GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UAAAAUAC 5392
3185 UUCCUGUA G GGGGCGAU 2184 AUCGCCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UACAGGAA 5-393
3238 UUCCUGUA G GGGGCGAU 2184 AUCGCCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UACAGGAA 5393
3275 UUCCUGUA G GGGGCGAU 2184 AUCGCCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG UACAGGAA 5393
3186 UCCUGUAG G GGGCGAUA 2185 UAUCGCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUACAGGA 5394
3239 UCCUGUAG G GGGCGAUA 2185 UAUCGCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUACAGGA 5394
3276 UCCUGUAG G GGGCGAUA 2185 UAUCGCCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CUACAGGA 5394
3187 CCUGUAGG G GGCGAUAU 2186 AUAUCGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUACAGG 5395
3240 CCUGUAGG G GGCGAUAU 2186 AUAUCGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUACAGG 5395
3188 CUGUAGGG G GCGAUAUA 2187 UAUAUCGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCUACAG 5396
3241 CUGUAGGG G GCGAUAUA 2187 UAUAUCGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCUACAG 5396
3277 CCUGUAGG G GGCGAUAA 2188 UUAUCGCC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCUACAGG 5397
3278 CUGUAGGG G GCGAUAAA 2189 UUUAUCGC GGA GCCGUUAGGC UCCCUUCAAGGA GCCGUUAGGC UCCGGG CCCUACAG 5398
Input Sequence = NM_001285. Cut Site = G/.
Arm Length = 8. Core Sequence = GGAGGAAACUCC CU UCAAGGACAUCGUCCGGG
Underlined region can be any X sequence or linker, as described herein.
NM_001285 (Homo sapiens chloride channel, calcium activated, 1 (CLCAl) mRNA, 3311 bp)
Table IX: Human CLCAl GeneBIoc and Target Sequence 249.021
Figure imgf000140_0001
Figure imgf000141_0001
lower case = 2OMe; A = riBo A Upper Case = DeoxyriBose (DNA) s = phosphorothioate linkages B = inverted aBasic U = 2'-C-allyI Uridme 6 = riBo G
Table X: PCR Primers 249.021
Figure imgf000142_0001

Claims

CLAIMS What is claimed is:
1. A nucleic acid molecule that down regulates expression of CLCAl (Chloride Channel Calcium Activated) gene.
2. The nucleic acid molecule of claim 1, wherein said nucleic acid molecule is used to treat conditions selected from the group consisting of Chronic Obstructive Pulmonary Disease (COPD), chronic bronchitis, asthma, cystic fibrosis, and obstructive bowel syndrome.
3. The nucleic acid molecule of claim 1, wherein said nucleic acid molecule is an enzymatic nucleic acid molecule.
4. The nucleic acid molecule of claim 3, wherein a binding arm of said enzymatic nucleic acid molecule comprise sequences complementary to any of sequences having SEQ ID NOs:l-2189 and 5399-5416.
5. The nucleic acid molecule of claim 3, wherein said enzymatic nucleic acid molecule comprises any of sequences havingSEQ ID NOs:2190-5398 and
5425-5434.
6. The nucleic acid molecule of claim 1, wherein said nucleic acid molecule is an antisense nucleic acid molecule.
7. The nucleic acid molecule of claim 6, wherein said antisense nucleic acid molecule comprises sequences complementary to any of sequences having
SEQ ID NOs:l-2189 and 5399-5416.
8. The nucleic acid molecule of claim 6, wherein said antisense nucleic acid molecule comprise any of sequences having SEQ ID NOs:5417-5424.
9. The nucleic acid molecule of claim 3, wherein said enzymatic nucleic acid molecule is in a hammerhead (HH) motif.
10. The nucleic acid molecule of claim 3, wherein said enzymatic nucleic acid molecule is in a hairpin, hepatitis Delta virus, group I intron, VS nucleic acid, amberzyme, zinzyme or RNAse P nucleic acid motif.
11. The nucleic acid molecule of claim 3, wherein said enzymatic nucleic acid molecule is in an Inozyme motif.
12. The nucleic acid molecule of claim 3, wherein said enzymatic nucleic acid molecule is in a G-cleaver motif.
13. The nucleic acid molecule of claim 3, wherein said enzymatic nucleic acid molecule is a DNAzyme.
14. The nucleic acid molecule of claims 3 or 6, wherein said nucleic acid molecule comprises between 12 and 100 bases complementary to RNA of a CLCAl gene.
15. The nucleic acid molecule of claims 3 or 6, wherein said nucleic acid molecule comprises between 14 and 24 bases complementary to RNA of a CLCAl gene.
16. The nucleic acid molecule of claim 1, wherein said nucleic acid molecule is chemically synthesized.
17. The nucleic acid molecule of claim 1, wherein said nucleic acid molecule comprises at least one 2 '-sugar modification.
18. The nucleic acid molecule of claim 1, wherein said nucleic acid molecule comprises at least one nucleic acid base modification.
19. The nucleic acid molecule of claim 1, wherein said nucleic acid molecule comprises at least one phosphate backbone modification.
20. A mammalian cell comprising the nucleic acid molecule of claim 1.
21. The mammalian cell of claim 20, wherein said mammalian cell is a human cell.
22. A method of reducing CLCAl activity in a cell, comprising the step of contacting said cell with the nucleic acid molecule of claim 1, under conditions suitable for said reduction of CLCAl activity.
23. A method of treatment of a patient having a condition associated with the level of CLCAl, comprising contacting cells of said patient with the nucleic acid molecule of claim 1, under conditions suitable for said treatment.
24. The method of claim 23 further comprising the use of one or more therapies under conditions suitable for said treatment.
25. A method of cleaving RNA of a CLCAl gene, comprising contacting the nucleic acid molecule of claim 3 with said RNA under conditions suitable for the cleavage of said RNA.
26. The method of claim 25, wherein said cleavage is carried out in the presence of a divalent cation.
27. The method of claim 26, wherein said divalent cation is Mg^+.
28. The nucleic acid molecule of claim 1, wherein said nucleic acid comprises a cap structure, wherein the cap st cture is at the 5'-end or 3'-end or both the
5 '-end and the 3' -end.
29. The enzymatic nucleic acid molecule of claim 9, wherein said hammerhead motif comprises sequences complementary to any of sequences having SEQ ID NOs: 1-575.
30. The enzymatic nucleic acid molecule of claim 11, wherein said Inozyme motif comprises sequences complementary to any of sequences having SEQ ID NOs.576-1210.
31. The enzymatic nucleic acid molecule of claim 12, wherein said G-cleaver motif comprises sequences complementary to any of sequences having SEQ ID NOs:1211-1429.
32. The enzymatic nucleic acid molecule of claim 13, wherein said DNAzyme comprises sequences complementary to any sequence shown as substrate sequences in Table VR.
33. The enzymatic nucleic acid molecule of claim 10, wherein said zinzyme comprises sequences complementary to any sequence shown as substrate sequences in Table VI.
34. The enzymatic nucleic acid molecule of claim 10, wherein said amberzyme comprises sequences complementary to any sequence shown as substrate sequences in Table VIE.
35. An expression vector comprising at least one nucleic acid molecule of claim 1, in a manner that allows expression of the nucleic acid molecule.
36. A mammalian cell comprising an expression vector of claim 35.
37. The mammalian cell of claim 36, wherein said mammalian cell is a human cell.
38. The expression vector of claim 35, wherein said nucleic acid molecule is an enzymatic nucleic acid molecule.
39. The expression vector of claim 35, wherein said expression vector further comprises an antisense nucleic acid molecule complementary to RNA of a CLCAl gene.
40. The expression vector of claim 35, wherein said expression vector comprises at least two said nucleic acid molecules.
41. The expression vector of claim 40, wherein the at least two nucleic acid molecules are the same.
42. The expression vector of claim 40, wherein the at least two nucleic acid molecules are different.
43. The expression vector of claim 40, wherein one said expression vector further comprises an antisense nucleic acid molecule complementary to RNA of a CLCAl gene.
44. The expression vector of claim 40, wherein one said expression vector further comprises an enzymatic nucleic acid molecule complementary to RNA of a CLCAl gene.
45. A method for treatment of chronic obstructive pulmonary disease comprising the step of administering to a patient the nucleic acid molecule of claim 1 under conditions suitable for said treatment.
46. A method for treatment of cystic fibrosis comprising the step of administering to a patient the nucleic acid molecule of claim 1 under conditions suitable for said treatment.
47. An enzymatic nucleic acid molecule that cleaves RNA derived from a CLCAl gene.
48. The enzymatic nucleic acid molecule of claim 47, wherein said enzymatic nucleic acid molecule is selected from the group consisting of Hammerhead, Hairpin, Inozyme, G-cleaver, DNAzyme, Amberzyme and Zinzyme.
49. The method of claims 45 or 46, wherein said method further comprises administering to said patient the nucleic acid molecule of claim 1 in conjunction with one or more other therapies.
50. The method of claim 49, wherein said other therapies are therapies selected from the group consisting of oxygen therapy, bronchodilators, corticosteroids, antibacterials, vaccinations, acetylcysteine, mucokinetic agents, and DNase (Pulmozyme) treatments.
51. The nucleic acid molecule of claim 9, wherein said nucleic acid molecule comprises at least five ribose residues, at least ten 2'-O-methyl modifications, and a 3'- end modification.
52. The nucleic acid molecule of claim 51, wherein said nucleic acid molecule further comprises a phosphorothioate core with both 3' and 5' -end modifications.
53. The nucleic acid molecule of claims 51 or 52, wherein said 3' and/or 5'- end modification is 3 '-3' inverted abasic moiety.
54. The nucleic acid molecule of claim 3, wherein said nucleic acid molecule comprises at least five ribose residues; at least ten 2'-0-methyl modifications, and a 3'- end modification.
55. The nucleic acid molecule of claim 54, wherein said nucleic acid molecule further comprises phosphorothioate linkages on at least three of the 5' terminal nucleotides.
56. The nucleic acid molecule of claim 54, wherein said 3'- end modification is 3 '-3' inverted abasic moiety.
57. The enzymatic nucleic acid molecule of claim 13, wherein said DNAzyme comprises at least ten 2'-O-methyl modifications.
58. The enzymatic nucleic acid molecule of claim 57, wherein said DNAzyme further comprises phosphorothioate linkages on at least three of the 5' terminal nucleotides.
59. The enzymatic nucleic acid molecule of claim 57, wherein said DNAzyme further comprises a 3 '-end modification.
60. The enzymatic nucleic acid molecule of claim 59, wherein said 3'- end modification is 3 '-3' inverted abasic moiety.
PCT/US2001/024970 2000-08-09 2001-08-09 Method and reagent for the inhibition of calcium activated chloride channel-1 (clca-1) WO2002011674A2 (en)

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1234878A1 (en) * 1999-11-24 2002-08-28 Takeda Chemical Industries, Ltd. Use of disease-associated gene
JP2007505162A (en) * 2003-05-28 2007-03-08 ワイス Soluble CLCA-1 and antagonists to CLCA-1
EP3263115A1 (en) * 2016-06-30 2018-01-03 Technische Hochschule Mittelhessen Invention relating to an agent combating pseudomonas aeruginosa

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1995011304A1 (en) * 1993-10-18 1995-04-27 Ribozyme Pharmaceuticals, Inc. Catalytic dna
WO1997026270A2 (en) * 1996-01-16 1997-07-24 Ribozyme Pharmaceuticals, Inc. Synthesis of methoxy nucleosides and enzymatic nucleic acid molecules
WO1998032846A2 (en) * 1997-01-23 1998-07-30 Ribozyme Pharmaceuticals, Inc. Enzymatic nucleic acid treatment of diseases or conditions related to levels of c-fos
WO1999055857A2 (en) * 1998-04-29 1999-11-04 Ribozyme Pharmaceuticals, Inc. Nucleoside triphosphates and their incorporation into ribozymes

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1995011304A1 (en) * 1993-10-18 1995-04-27 Ribozyme Pharmaceuticals, Inc. Catalytic dna
WO1997026270A2 (en) * 1996-01-16 1997-07-24 Ribozyme Pharmaceuticals, Inc. Synthesis of methoxy nucleosides and enzymatic nucleic acid molecules
WO1998032846A2 (en) * 1997-01-23 1998-07-30 Ribozyme Pharmaceuticals, Inc. Enzymatic nucleic acid treatment of diseases or conditions related to levels of c-fos
WO1999055857A2 (en) * 1998-04-29 1999-11-04 Ribozyme Pharmaceuticals, Inc. Nucleoside triphosphates and their incorporation into ribozymes

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
COUTURE L A ET AL: "Anti-gene therapy: the use of ribozymes to inhibit gene function" TRENDS IN GENETICS, ELSEVIER SCIENCE PUBLISHERS B.V. AMSTERDAM, NL, vol. 12, no. 12, 1 December 1996 (1996-12-01), page 509 XP004071051 ISSN: 0168-9525 *
GRUBER A D ET AL: "GENOMIC CLONING, MOLECULAR CHARACTERIZATION, AND FUNCTIONAL ANALYSIS OF HUMAN CLCA1, THE FIRST HUMAN MEMBER OF THE FAMILY OF CA2+-ACTIVATED CI CHANNEL PROTEINS" GENOMICS, ACADEMIC PRESS, SAN DIEGO, US, vol. 54, 1998, pages 200-214, XP002945247 ISSN: 0888-7543 -& DATABASE GENBANK [Online] 14 December 1998 (1998-12-14) "Homo sapiens calcium-dependent chloride channel-1 (hCLCA1) mRNA, complete cds" Database accession no. AF039400 XP002219572 *
LUDWIG J ET AL: "Extending the cleavage rules for the hammerhead ribozyme: mutating adenosine15.1 to inosine15.1 changes the cleavage site specificity from N16.2U16.1H17 to N16.2C16.1H17" NUCLEIC ACIDS RESEARCH, OXFORD UNIVERSITY PRESS, SURREY, GB, vol. 26, no. 10, 15 May 1998 (1998-05-15), pages 2279-2285, XP002084035 ISSN: 0305-1048 *
VAISH ET AL: "In vitro selection of a purine nucleotide-specific hammerhead-like ribozyme" PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF USA, NATIONAL ACADEMY OF SCIENCE. WASHINGTON, US, vol. 95, no. 5, 1 March 1998 (1998-03-01), pages 2158-2162, XP002097465 ISSN: 0027-8424 *

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1234878A1 (en) * 1999-11-24 2002-08-28 Takeda Chemical Industries, Ltd. Use of disease-associated gene
EP1234878A4 (en) * 1999-11-24 2003-06-11 Takeda Chemical Industries Ltd USE OF A GENES RELATED TO A DISEASE
JP2007505162A (en) * 2003-05-28 2007-03-08 ワイス Soluble CLCA-1 and antagonists to CLCA-1
EP3263115A1 (en) * 2016-06-30 2018-01-03 Technische Hochschule Mittelhessen Invention relating to an agent combating pseudomonas aeruginosa

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