JP2002300884A - Anti-sense oligonucleotide having inhibitory action on drebrin a expression - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide an anti-sense oligonucleotide having promising usefulness as a therapeutic drug for fragile X syndrome or the like and capable of specifically inhibiting drebrin A expression. SOLUTION: An anti-sense oligonucleotide capable of hybridizing with 24-mer S-oligo(5'-AGGAAGGCCCACTGTCCGATGCCT-3') being an anti-sense chain of aggcatcggacagtgggccttcct (sequence number 5) in ins2 region which specifically exists in drebrin A and the sequence is suitably maintained beyond animal species, and the base sequence represented by the sequence number 5 under a stringent condition and having an inhibitory action on drebrin A expression, and derivatives thereof are produced.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to an antisense oligonucleotide having an inhibitory action on drebrin A expression, a derivative thereof, a vector carrying them, a therapeutic drug for a disease caused by overexpression and hyperfunction of drebrin A, and the like.

[0002]

2. Description of the Related Art Nerve cells have a very complicated shape that cannot be found in other somatic cells. The projections are extended. Dendritic dendrites are spines
It has a myriad of spine structures called e), which form the postsynapse, which has the function of receiving information from other cells.
This neuron-specific morphology is determined by the nerve-specific actin binding protein. On the other hand, the present inventors have discovered the world's first actin-binding protein, Drebrin, which is abundantly expressed in developing neurons (J. Neur
ochem. 44, 1210-1216 (1985), J. Biochem. 117, 231-2
36 (1995)) that the drebrin is involved in neuronal morphogenesis, particularly projection formation, by altering the properties of actin fibers (J. Neurosci. Res. 38: 149-).
159 (1994), Exp. Cell Res. 215: 145-153 (1994),
J. Biol. Chem. 269: 29928-29933 (1994)) and in developing and migrating neurons, it is present throughout the cell body and processes, but in mature neurons it is specifically located in the spine structure. Existence (J. Neurosci. 15: 7161-7170 (1996), Dev.B
rain Res. 29, 233-244 (1986), Brain Res. 413, 374-
378 (1987)). Drebrin includes embryonic type drebrin E and adult type (adul)
t type) and two isoforms of drebrin A (J. Biochem. 117, 231-236 (1995)).
Drebrin A, which is specifically found in the spine of mature neurons, has the characteristic of being expressed only in neurons (Dev. Brain Res. 29, 233-244 (1986), Brain).
Res. 413, 374-378 (1987)). Drebrin A is alternat
A sequence in which a region called "ins2" is added to drebrin E by an ive splicing mechanism. That is,
Gtc from 95 'base to 956 base to 1093 base
gtccgtactgcccttttcataaaaggc
atcggacaggggggccttcctcctcc
tcctctttcctcctcttcccctccac
ggactcccttttccctatatcacctg
cccaccgcaccccaaacctctctttcc
tccctcccat) [SEQ ID NO: 1] was inserted and expressed (Mol. Brain Res. 19, 101-114 (199)
3), Neuroreport 3, 109-112 (1992)).

Recently, the present inventors have found that when primary cells of drebrin A are expressed in primary cultured neurons, they are automatically collected on dendritic spines, and the length of the spines is increased, and the amount of one protein synthesized is changed. Report that spine morphology can be altered by causing
Neurosci. 19, 3918-3925 (1999)). In addition, regarding the protein family to which drebrin belongs, it has been suggested that drebrin may be classified as a type of actin-binding protein having an ADF Homology Domain (Mol. B
iol. Cell 9, 1951-1959 (1998)). SH3P7 was discovered as a homologue of Drebrin,
The function of H3P7 is being elucidated (Mol. Cell. B
iol. 19, 1539-1546 (1999), Nature Biotech. 14, 74
1-744 (1996)).

[0004] In addition, the neurotrophic factor BDNF (Brain-de
(Ribbed neurotrophic factor) to produce exogenous B
DNF blocks estradiol's effect of increasing spine density in vivo and in vitro, and suppression of BDNF expression using selective antisense oligonucleotides has an effect similar to estradiol, indicating that anti-BDN
The role of BDNF involved in the regulation of spine density has been elucidated, including the increase in spine density by the F antibody (Proc. Natl. Acad. Sci. USA 95, 11412-11417, 19).
98). In addition, fragile X syndrome (fragile X syndrome)
The most frequent cause of hereditary mental retardation caused by the presence of fragile sites on the X chromosome is abnormal expression of the gene FMR1 on the X chromosome, mainly caused by defective expression, and abnormal expression of FMR1 Is accompanied by morphological abnormalities of the cranial nervous system, but there is currently no cure for cranial nerve diseases such as fragile X syndrome.

[0005]

An object of the present invention is to provide an antisense oligonucleotide which can be expected to be useful as a therapeutic agent for fragile X syndrome and which can specifically suppress the expression of drebrin A. It is in.

[0006]

Means for Solving the Problems The present inventors were the first in the world to discover and request an actin-binding protein, drebrin, which is abundantly expressed in developing neurons, and conducted a comprehensive and multifaceted study on drebrin. I have. Recently, the present inventors have determined that the genomic gene for mouse drebrin A (Dbn
1) was isolated for the first time in the world and its sequence was determined. The exon-intron configuration of mouse Dbn1 consisting of the nucleotide sequence shown in SEQ ID NO: 2 is shown in FIG. From the chromosome map shown in FIG. 2, mouse Dbn1 having a total length of 2067 bp, in which the first exon starts from the 6767th position, is located almost at the center of chromosome 13 and is close to interleukin-
9 (Il9) and paired-like homeodomein transcription f
You can see that actor (Pitx1) exists.

[0007] The agreement between the mouse drebrin A cDNA (accession number: AF187147) and the rat drebrin A cDNA shown in SEQ ID NO: 3 is 94.07%. The agreement rate at the amino acid level with rat drebrin A is 9
Although it is 5.35%, the mouse ins2 sequence constituting exon 11 is completely (100%) identical to the rat ins2 sequence.
Was matched. The present inventors have focused on the ins2 region, which is not present in drebrin E but specifically present in drebrin A, and whose sequence is well conserved across animal species. An experiment was carried out to prepare a chain and specifically suppress the expression of drebrin A in neurons.
Agg from 980b to 1003b from the 5 'side of NA
The 24-mer S which is the antisense strand of the region of catcggacaggtgggccttcct [SEQ ID NO: 5]
-Oligo (5'-AGGAAGGCCCACTGTCC
GATGCCT-3 ') [SEQ ID NO: 6] is Drebrin A
Have been found to significantly reduce the expression level of, resulting in the completion of the present invention.

That is, the present invention relates to an antisense oligonucleotide comprising the nucleotide sequence of SEQ ID NO: 6 (claim 1) and a nucleotide sequence which hybridizes with the nucleotide sequence of SEQ ID NO: 5 under stringent conditions, and A antisense oligonucleotide having an A expression inhibitory action (Claim 2); a derivative of the antisense oligonucleotide according to Claim 1 or 2 (Claim 3);
Or a vector carrying the antisense oligonucleotide according to claim 2 and / or the derivative of the antisense oligonucleotide according to claim 3 (claim 4), or an antibody that specifically recognizes a peptide having the amino acid sequence represented by SEQ ID NO: 7 (Claim 5) or an antibody according to claim 5, which is a monoclonal antibody (claim 6), an antisense oligonucleotide according to claim 1 or 2, and / or an antisense according to claim 3. A therapeutic agent for a disease caused by overexpression and hyperfunction of drebrin A containing an oligonucleotide derivative as an active ingredient (Claim 7), the antisense oligonucleotide according to Claim 1 or 2, and / or
Alternatively, the present invention relates to a therapeutic agent for a disease caused by overexpression and hyperfunction of drebrin A, comprising a derivative of the antisense oligonucleotide according to claim 3 and a pharmaceutically acceptable reagent for introduction into a cell (claim 8).

[0009]

BEST MODE FOR CARRYING OUT THE INVENTION As the antisense oligonucleotide of the present invention, a nucleotide sequence represented by SEQ ID NO: 6 (5'-AGGAAGGCCCCACTGTCCGATG
CCT-3 ') or the base sequence shown in SEQ ID NO: 5 (aggcatcggacaggtgggcc).
ttcct) and any other antisense oligonucleotide (antisense DNA or antisense RNA) that hybridizes under stringent conditions and has the effect of suppressing drebrin A expression. Antisense DNA is preferred because of its higher stability. The antisense oligonucleotide of the present invention can remarkably suppress the expression of drebrin A, and can also be used as a probe because of its selective hybridizing property, and can be used as a probe from other vertebrate cDNA libraries. A cDNA that is a homologous gene of the drebrin A gene in the animal that hybridizes with the probe can also be obtained.

The conditions for hybridization under the above stringent conditions include, for example, 42
Hybridization at 1 ° C. and 1 × SSC, 0.
Washing treatment at 42 ° C. with a buffer containing 1% SDS, hybridization at 65 ° C., and 0.1 ×
Washing at 65 ° C. with a buffer containing SSC and 0.1% SDS can be mentioned. The factors that affect the stringency of hybridization include various factors other than the above-mentioned temperature conditions, and those skilled in the art can appropriately combine various components and have the same properties as the above-described stringency of hybridization. Stringency can be realized. The antisense oligonucleotide obtained in this manner has 12 to 39 bases, particularly 15 to 25 bases.
Antisense DNA or antisense RNA of bases or less
Is preferred.

The method for synthesizing the antisense oligonucleotide of the present invention is not particularly limited.
NA synthesizer (“381” manufactured by Applied Biosystems Inc.)
A ") and DNA / RNA synthesizer (Applied Biosystems I
394, manufactured by nc., etc.), a phosphoramidite method, a phosphorothioate method, a phosphotriester method, and the like using a general synthesizer. In this case, the target antisense polynucleotide or the target antisense polynucleotide can be obtained by performing the operation according to the instructions attached to the DNA synthesizer or the like, and purifying the resulting synthetic product by HPLC using reverse phase chromatography or the like. Derivatives can be obtained.

In order to improve the stability and affinity of the antisense oligonucleotide of the present invention for cells,
As long as its activity is not significantly reduced, it can be used as a derivative in which the hydroxyl group of the phosphate ester group or ribose moiety is substituted with another stable group. Specific examples of the derivative of such an antisense oligonucleotide include those in which a phosphate group is substituted with a thiophosphate group or a methylphosphonate group, and a hydroxyl group of a ribose moiety is an alkoxy group such as methoxy or allyloxy, an amino group, Alternatively, those substituted with a fluorine atom or the like can be given.

[0013] The antisense oligonucleotides and derivatives thereof of the present invention suppress the expression of drebrin A in nerve cells, and are considered to be one of the causes of overexpression and hyperfunction of drebrin A protein in nerve cells. There is a great possibility that certain diseases, such as fragile X syndrome (fragile X syndrome), can be effectively treated. When treating a disease that is considered to be one of the causes of overexpression and hyperfunction of the drebrin A protein, the antisense oligonucleotide of the present invention or a derivative thereof 0.01 to 100 μM, preferably 0.1 to 100 μM.
10 μM can be administered, and if necessary, a pharmaceutically acceptable intracellular introduction reagent such as a lipofectin reagent, a lipofectamine reagent, a DOTAP reagent, T
fx reagent, artificial synthetic lipid vesicle, liposome, membrane fusion reagent, polymeric micelle-forming reagent, polymer carrier, or other intracellular introduction reagent. In this case, the antisense oligonucleotide of the present invention can be administered alone, but a pharmaceutically acceptable ordinary carrier, binder, stabilizer, excipient, diluent, pH buffer,
Various compounding ingredients for preparations such as disintegrants, solubilizers, solubilizers, isotonic agents and the like can be added. Such therapeutic agents can be administered orally or parenterally. That is, it can be orally administered in a commonly used dosage form, for example, a powder, granule, capsule, syrup, suspension or the like, or, for example, a solution, emulsion, suspension or the like. Can be parenterally administered topically in the form of injections. For example, in the case of an injection, it can be prepared by dissolving the antisense oligonucleotide of the present invention in water, physiological saline, a glucose solution, or the like,
If necessary, a buffer, a preservative, a stabilizer or the like may be contained. The dosage may vary depending on the degree of symptoms of each disease, the administration method, etc., but as a guide, for intravenous administration, 1 mg / kg to 1 g / kg, preferably 5 mg / kg to 500 mg / kg per body weight is exemplified. can do.

In addition, plasmids and viral vectors designed to express the antisense oligonucleotide sequence of the present invention are used for gene therapy in order to promote their uptake into cells and to enhance their directivity to target cells. It can also be used as a vector. Such vectors include herpes virus (HSV) vectors, adenovirus vectors, human immunodeficiency virus (HI)
V) A virus vector such as a vector can be preferably mentioned, and among these virus vectors, an HSV vector is preferable. HSV vectors have high neurotrophic properties, are safe because HSV is not integrated into chromosomal DNA of cells, and can regulate the expression period of transgenes. When a viral vector is used, the use of the inverted repeat sequence recognized by the recombinase, for example, the loxP sequence derived from Escherichia coli P1 phage or the wild-type FRT sequence derived from yeast Saccharomyces cerevisiae, allows the antisense oligonucleotide of the present invention to be obtained at a desired time. The nucleotide can be expressed.

The antibody of the present invention has a nucleotide sequence represented by SEQ ID NO: 5 (aggcatcggacaggtgggcctt).
The antibody of the present invention is not particularly limited as long as it is an antibody that specifically recognizes KASDSGPSS [SEQ ID NO: 7], which is a translation peptide of cct), or a region containing this peptide. Monoclonal antibodies, polyclonal antibodies Specific examples thereof include immunospecific antibodies such as chimeric antibodies, single-chain antibodies, and humanized antibodies. These can be prepared by a conventional method using the above-mentioned KADSDSGPSS as an antigen according to a conventional protocol. ,
Among them, a monoclonal antibody is more preferable in terms of its specificity. KASDSG such as the monoclonal antibody
Antibodies that specifically bind to PSS are considered to be one of the causes, for example, in diagnosing diseases caused by mutation or deletion of drebrin A, and overexpression and hyperfunction of drebrin A protein in nerve cells. It is useful in the treatment of the diseases in which

[0016]

EXAMPLES Hereinafter, the present invention will be described more specifically with reference to examples, but the scope of the present invention is not limited to these examples. Example 1 (Culture of nerve cells) Sprague-Dawley rats on gestation day 20 were anesthetized with ether,
After decapitation, the fetus was removed. The cerebral cortex was removed, the meninges were peeled off, put in a CGBD solution containing papain, and left in a thermostat at 37 ° C. for 15 minutes. The supernatant was removed by suction, washed with 5 ml of MEM medium, and the supernatant was removed again by suction.
Add 3 ml of medium and 2 ml of horse serum and pipette.
The nerve cell-containing solution is filtered and centrifuged, the supernatant is discarded, and the mixture is stirred in a MEM medium containing 5% bovine serum and 5% horse serum, and placed in a dish having a diameter of 3.5 cm.
⁶ / 2ml (for western blot) or 2.0 × 1
At a concentration of 0 ^6/2 ml (for immunostaining) were respectively injected by 2 ml. The nerve cells in these dishes were cultured at 37 ° C. in a CO ₂ incubator. From the fifth day, A
Using a glial conditioned medium containing 5 μM of raC, the culture was continued twice a week by changing the medium in half each.

Example 2 (Administration of Antisense Oligonucleotides) On the 12th day from the start of the culture, the following four types of antisense oligonucleotides were administered to the medium to a final concentration of 10 μM. Cells were collected and subjected to immunostaining and Western blot. 5'-TTATGAAAGGGCAGTACGGACGAC-3 '[SEQ ID NO: 8] 5'-AGGAAGGCCCACTGTCCGATGCCT-3' [SEQ ID NO: 6] 5'-TATAGGGAAAGGGAGTCCGTGGAG-3 '[SEQ ID NO: 9] 5'-GGTTTGGGGTGCGGTGGCAGGTGA-3' [SEQ ID NO: 10] As a positive control, four kinds of reversed antisense strands in which the sequence of the antisense strand was reversed were used.

Example 3 (Western blot analysis) Western blot analysis was performed to examine the expression level of drebrin A protein in nerve cells two days after administration of the antisense chain. The extract of nerve cells was separated by SDS-polyacrylamide gel electrophoresis, and the separated proteins were separated by a membrane filter ("Immobi" manufactured by MILIPORE).
lon transfer membranes "). The blotted membrane was washed with T containing 5% skim milk.
Anti-Drebulin A antiserum diluted 5 times with BS (Exp. Cell
Res. 215, 145-153 (1994); anti-drebrin A polyclonal antibody) containing solution for 60 minutes at room temperature. After washing with TBS containing 0.05% Tween 20, HRP diluted with TBS containing 5% skim milk
The above-mentioned incubated membrane was immersed in a solution of F (ab ') ₂ fragment (manufactured by Kappel) of a labeled anti-rabbit IgG antibody. The chemiluminescence generated by the antigen-specific HRP reaction was measured using Kodak Scientific Imaging Film (X-OMA).
TAR, Kodak) and ECL detection kit (Amersham Pharmac)
ia Biotech). As a result, agg was placed at 980 base to 1003 base from the 5 ′ side.
antisense strand 5'-AGGAAGGCCCAC for the region of catcggacagggggcctccct
In the case of TGTCCGATGCCT-3 'administration, it was found that it was effective in specifically lowering the expression level of drebrin A, as compared with the case of the control to which no antisense strand was added and the case of the positive control to which the reversed antisense strand was administered. (FIG. 3), however, the expression of drebrin A was not suppressed in the other three types of antisense strands.

Example 4 [Immunostaining] Antisense strand 5'-AGGAAGGCCCACTGT
Neurons 2 days after CCGATGCCT-3 'administration were treated with PBS supplemented with 3.5% paraformaldehyde.
After shaking and fixing at 4 ° C. for 15 minutes, the plate was washed in TBS at room temperature for 5 minutes, blocked in PBS containing 3% BSA at room temperature for 10 minutes, and subjected to anti-drebrin A antiserum (Exp. Cell Res. 215, 145-153 (1994); anti-drebulin A polyclonal antibody) was reacted with a solution prepared by diluting with PBS containing 3% BSA at room temperature for 60 minutes. The reacted specimens were washed by shaking three times in PBS at room temperature for 5 minutes, and then washed with PB containing 3% BSA.
Staining was performed at room temperature for 30 minutes using a FITC-labeled anti-mouse IgG antibody (manufactured by Kappel) diluted 100-fold with S. Keep these stains in PBS at room temperature for 5
After shaking for 3 minutes x 3 minutes for washing, the cells were sealed with Permaflow (manufactured by Shandon) and observed with a fluorescence microscope. The result is shown in FIG.

When the distribution of drebrin A in the cells by immunostaining was examined in comparison with the neuronal morphology by DiI staining, in the normally developing nerve cells, spine, which is a spine structure of dendrites, was found. A is located in dendrites, not spines, and is located in the same manner as immature nerve cells, suggesting a delayed development of the nervous system. Furthermore, 40 to 63 spines were counted per neuron, and the spine length of each neuron was measured. FIG. 5 shows the results. From FIG. 5, the antisense strand 5 '
-AGGAAGGCCCACTGTCCGATGCCT
In the −3′-administered group (AS), the spine length was statistically significantly higher than in the control antisense-strand-free group (Cont) and the positive control reversed-sense-chain-administered group (RAS). It was getting shorter. The spine density per dendrite was also measured. FIG. 6 shows the results. As can be seen from FIG. 6, the spine density was measured for the antisense strand 5′-AGGAAG.
To find a statistically significant difference between the GCCCACTGTCCGATGCCT-3'-administered group (AS) and the control antisense-strand-free group (Cont) or the positive control reversed-sense-administered group (RAS). Could not.

[0021]

When the antisense oligonucleotide of the present invention is used, the expression level of drebrin A protein is remarkably reduced, and in a nerve cell in which the expression level of drebrin A is reduced, it is an important structure in the nerve cell. Since the length of the spine structure of the dendrites is shortened, it is possible to regulate the nerve synaptic function, and it can be applied to the treatment of psychiatric / neurological diseases and nerve damage.

[0022]

[Sequence List] SEQUENCE LISTING <110> JAPAN SCIENCE AND TECHNOLOGY CORPORATION <120> Antisense oligonucleotide with inhibitory action of Drebrin A expression <130> A091P20 <140> <141> <160> 10 <170> PatentIn Ver. 2.1 <210> 1 <211> 138 <212> DNA <213> Rattus norvegicus <400> 1 gtcgtccgta ctgccctttc ataaaggcat cggacagtgg gccttcctcc tcctcctctt 60 cctcctcttc ccctccacgg actccctttc cctatatcac ctgccaccgc accccaaacc 120 tctcttcctc cctcccat 138 <210> 2 <211> 20674 <212> DNA <213> Mus musculus <400> 2 gatcaccctg ttccggagct ggagtgtgag gctctatggc tggaacccac acttggaaat 60 gaatgtcttt atgccaggca ggggaggcca ttagcaccct tagagagatt gcagggcccc 120 cagccaacat ggcagagtcc ctagtagggg tctgtgtggg cagcaggtga gggaggggct 180 tgttcctgct ttgagcagct tggacttctt tgtagtcagc agggagaggt ggagggtagg 240 ggaccacttc tccagaactg aggcttctgt gctgcctcct ctggtctcct cagacctctc 300 cacgtggcct gggtacacac agaaggcaca cccgcctctc caagccttgc tacaggaaag 360 gagtgtggag gcagcaggct aaaggccctg gtggctgccc accagggagc agcaggtagc 420 tgagacccag agccatcctt gagaatgttg gaagggaccc cttcccaatg tgaggaaaga 480 ggcttaagct agattccagg gtcattggcc attgtagtgg gaagagatgc ttattggatc 540 agagccaggg gagaccttgg gagacctcct ttaagtgaga aatggggata attttccaga 600 agtactaggt ggcttcagcc atggaacact ggatagaggt gatagaggta tcctctttcc 660 cttagccttg cctcctcatg gggatccatc ccaggagcac accctgttat ggccaaaaga 720 cgctccattg gactgacttg ggtcacacac aggtccatcc gtgatgtcac aggtttgacc 780 cacaaggcct ggattacagg gtacaggaag aatcagaaga tacaaacagg aaggctgagg 840 ataatcgagc ac aaatgtcc agtgtggttc agccctccac ggtggggcct ctgctcctgg 900 actccatgcc cagacctctc agcctagctt ccagaaaggg tgaccaaagt ttaggctgtt 960 gagtccataa tcgaagaaat tcctgtgctg ctggtgtgtg tgtgtgtgtg tgtgtgtgtg 1020 tgtgtaagag agaggagaga tagagaggta gagaaagata agagaaagag gctacttagc 1080 tatgtataac ttagggagtt gttttgtgtg tgtgtgtgtg agagagagtg tgtatgtgtg 1140 tgtaagagag agagagacag agatagagag agataagaga gagaggctac ttagctatgt 1200 ataacttggg gagttgtttt gtgtgtgtgt gtgagagaga gagagagaga atgaatgtgt 1260 gtgtaagaga gagagagaga gagagagaga gagagagaga gagagagaga acgggacttt 1320 accactcagc tgtgtatcta ggtgagttct tgtctggccc ctaacccatc cagtgtacta 1380 cttgtgtgtt ttgctacttt aaacaattcc tagatacttt ggggggggga tcatcaaatt 1440 aaccacacct gatgtcccag agtgactgag gcaggaggat cacaaattca aagtttgcct 1500 gggctacaca gtgagtttcg ggccactttg ggcaacttag tgagaccttg tcttgaaata 1560 aaaagggtgc taaagatata gctcagatct accagatcac ttgtagacaa aaaacgtgtg 1620 gagcgttttc ttgattaata attgatgtgg gaggacccaa ctcactgaag gcggtgacac 1680 cccttggtag atgatcctg g ggtgaataac aaatcaggct gagcaagtcg gagcaagcca 1740 gcgagcagca ctcctctgtg gtctctgcct cagttccagc ctacaggttt ctgccctgac 1800 ttccctccag aatggactat aaactggtag gctaaaataa acccttcctt cccaagatgc 1860 tttgggtcat ggtgttttag cacagcaaca gaaagcccac taagacaaat catcaaaccg 1920 aagccaggca cacaggtaat ctctgctgag atgggcattt gagatacaga tcctgcctca 1980 ctccccgccc ccatctaaag tcacaccaat gacaaggcac agttgtgtga ggatagatac 2040 atttgttgca agaagcagag ctaatgtact tggcccccag tgccaaggtc catgcctggg 2100 gactgctgag cttggtattg gtgaggaggt acttttgaag gtgtactagg taggatctta 2160 agggtcagga tgaggccagg gaaggctctg ccagctgata ctgaaataat cccgcagccc 2220 cacatactcc cctcagtctt gaaacctatg aaaggtgtgg tggaacacag agctggaagc 2280 ttggggagga cagggctgcc aggaggaaag ggtggaggtg ggggtgggga gctccagtgt 2340 tctaagttac catccaggcc cattttatac tcccaacaac agggtctcaa aaggaaactc 2400 agaaggggga gcgctttagc cgaagacaca cagtaaggcg ctagtttctg aatctattct 2460 cttctgttac attctaggga cccaaaagcc atggggccag cctctcaatg tgtgggggga 2520 gctctgaggg tgggagagac tggc atgtca gtgatttgca gggtgatcaa tgccaggggg 2580 tggcagacac gaagcctggt gggaggacct ccaaggagac acctacgtgc cagccaccca 2640 ccacagccgg gtgtggtttt taggggctgc aaggaggcag ggtctctgag gagtgagcag 2700 tggccatctt ccttgacgac aagtacagcc aaggaaagag ctaaaataga gcaactgagc 2760 ctggccattt aaagatgatg acgttggcca aaaaggggcc aagaagcacg gaaataagaa 2820 aagcccaggg cttgtataag cctcagagcc cgccatccct taaaagtaaa acaggatgtg 2880 aggctgtggg atctgagcat cccttacgaa tgttgttgtt gttgttgttc ttgtttttct 2940 tccctttggt gctaagggtt tgacattggc tagagaggag ctcttaagct cacccagacg 3000 ctcgcagccc cagttctaaa atggtttcat ctctgggttt cttgtttgtt ttgtttttga 3060 gacagggccc ctctgtgtag ctctggctat cctggagctt cataagtaga ccaggctggc 3120 ctcaaatgca caaagatctg cttgcctctg cctccagaag gatcctgtcc tggtcctggc 3180 ccttcaccac ctctacacct ggttgtttgg cttggttttt tggaacagca gctcttggaa 3240 cccttggaga tggagaatgc tttctgcctc atctctgaat gctcggagcc cagccatcat 3300 catgctcgcc aagctgaaac cattctaacc cagccaccac atcgggtggc ttatgcctgt 3360 aatgcccgaa ttccgacccg acccggagtt tgagggcagc ttgaactaca gtgtgagctc 3420 tgtctcaaat aacaacagaa actaaacaag ggccagtggg gtggctcagt ggatatagcc 3480 agatggtcca tcgctggtgg gaggagagac cagactcttg gaagatgtct tctgacttgc 3540 ataggaagga aggctgaccc catgggatgc ttgtacccac acacatgcac aatgataatt 3600 aaaaaaaaat ggggggggga gggaagccca gcagaagagg gagcatgtgt gtgcgtgtgt 3660 gtgagagagt gtgtgtgtgt ctctgtgtgt ggcaggagag gggagaggag gaataataca 3720 aaaccaatgt cccatgatgt ggtgaagccc ggtaccctgt atgctaatct aaaaattaaa 3780 aaggagaaag aacttggatt tttatttcct ttgagacaaa gtctcacaat ataagcttgg 3840 ctgagctgga gctcacagag aaagctgctt gcctcttttt ccccagggct gggttggcac 3900 atgccatgac acccagacaa ggtttatcta tttatctatc tatctatcta tctatctatc 3960 tagatattta ttttatttgt ttttttttaa agatttattt atttattata tgtaagtaca 4020 ctgtagctgt cttcagacac tccagaagag ggcgtcaaat cttgttacag atggttgtga 4080 gccaccatgt ggttgctggg atttgaactt cagacctttg gaagagcagt cgggtgctct 4140 tacccactga gccatctcac cagccccttt atttatttta agatttactt atttaagccg 4200 ggcatggtgg cgcatgcctc taatcccagc actca ggagg caaaggcagg cggatttctg 4260 agttccaggc cagcctggtc tacaaaatga gttccaggac agccaggcta cacagagaaa 4320 ccctgtctcc aaaaaaaaaa aaaaaaaaaa aaaaaatttt tacttattta atgtatatga 4380 gtacactgtt gctgtcttca gacacactag aagagggcgt cagattccat tacagatggt 4440 tgtgagccac catgtggttg ctgggaattg aactcaggac ctttggaaga gcagccagtg 4500 ctctttaccg ctgagcgaaa tagaatgtcg tttttaaaaa taacgaaaac agcactggag 4560 agatggctta gcggttaaga gcactaattg ctcttctaga ggtcctgagt tcaattccca 4620 gcaaccataa ggtggctcac aagcatctgt aatgggatcc agtgccctct tctggtgtgt 4680 ctgaagacag caacagtata ctcatataca taaattaaaa agacaaaaac aaaaataaaa 4740 tgaaaacaga acaaaaaaat aatgcagatg ggaggtgaac tgggcgtggc tagagacagg 4800 tgctaggcaa ttggtgggca gacaggaatc catggagaca cttccaaggt gcacttaggt 4860 ctgctttttg cctatgaaat cgactaagta ggaggttagg agacatcaag ggtggggaag 4920 tagcaggaag gagagatgct gaagcctagt cctgacccca accgccaact cccctctgcc 4980 caggcgagca ctctagatac cgagaactgc agctctgttc ttggattttt ttggggcggg 5040 gggtttcgag acagggtttc tctgtgtagc cctggctgtc ctggaactca ctctgtagac 5100 caggctagcc tcgaactcag aaatctgcct gtctctgcct cctgagtgct gggatcaaag 5160 gcgtgcgcca ccactgcctg gcttgcagct ctgttctcac cttggcttcc tgctcctttc 5220 cttccccatc tgatgtcatc tctattcccc tctgccctaa ccattgagtt ccagctgcat 5280 ctgggatcag ggcccaggac tgaccactca ccaggactca ccataggttg gtggagaaac 5340 tcccaaggat ggggtgcatg ggttcaatgg gtacccatag ctcctgaagc agggttcttg 5400 tgaagtacag aggtttaagg ccaccctctc ccgtgtccca ttccaaggca ttgcagtgaa 5460 tgtaagtatt ggtttcttgt tggagatgat tgaccaagtc agagcaggaa ttcgacgaaa 5520 acagccacct cctgttcaca gatcagcccc aggggaaact tgctgaggca ctgttgcctg 5580 cctgtctgcc tgcctgcctg cctgcctgcc tgcctgcctg cctgcctggg aaatgggttg 5640 ggatgtccag tgggtatgga aaggttaacc agagctgtgg agggcttcag tgttctccat 5700 tccacaggaa acagtgcctg gaagctctgt gcagccatgt ccccatagag aagtctctga 5760 ggcccagtcc cttctgcact cagagctttg ggaaataaaa agtgccagaa agttctccct 5820 ggaatggctt tggaggagcc ctgccttggg gaccctactt gggcttcctt aggagggatg 5880 gtacttggaa aagaaggcat catttttttt tttttttttt tttga taccc atctgggagg 5940 aagaaaatga ctgaaagtaa aggatgaggg aggtggggtg taaacggctt gagacaaggt 6000 atggctaaga agaggggtct gagtggaatt cagacaggct ggcatagcaa gagggtatgg 6060 tgactaggtg ggatgtgtgc agaggtaagc atgggcctgt agctgagatg gaagtagtgg 6120 tggtagacta gggagggtcc tttcagagga aatggactac acgtaaccca ggcctggtct 6180 gatgaggaac tgaatccggg cagggacaga gtaaggacca taaggggttt aaggtagggt 6240 gtaacggacc caggaatgat gtgagtctgg gctggagtta aatggatcaa gggtgttggg 6300 ctgggtcaat tactctgggg agatggagag ggatgaatct gggcctggaa tcttggtggg 6360 tttatggcta gggaagaggg gtgtgctgcc aggttgggga cgaggacaag acaggtgcat 6420 gggactggcc ctgggagcag agcggaagct cagtgtccgg tgctggccag cagggtgagg 6480 ggcgtgtgtg ggacgccccg gggcaaactg ggaagggggg gcgggcctcc ccctgggccg 6540 ggccggggcg gcgctaggac cgagcgagcc gagggagtga gccgggcgga gccagcgccc 6600 cccgcccccc gccggccggc tcccctcccc cggccgctgg ctcgctcgct ctcagcgctg 6660 cagaggctct gaggcggcgg cggcgactcc ctcatcccct ccctccggtg tcggtcggtc 6720 cgcctgtgcg tgcgtctgtc cgttcggcct cggtccggcc cgcagcatgg ccggcgtcag 6780 cttcagcggc caccgcctgg agctgctggc ggcgtacgag gaggtgatcc gggaggagag 6840 cgcagccgac tggtgagccc cccgcccccg cccccgcccc cggccccttt gtccccaggc 6900 cgccgcgcgc ctttgtttcc ccgaggctct gccaccggga agggaggggg agaccccgaa 6960 atggtgcaac ggctcgggag agccggggag gggctgcctt tgtgcgagcc gggggagggg 7020 ccgctcgggc caggctgggc gcgcgggccg gggcgccctg aggccgccct aaggctcact 7080 gggggcgcgg gaccggacgc cagccacgcg ggtgtggagc ccagacgtgg ggcctggggt 7140 cctacggtcc tgtcggagcc catccccctt ctcaccctgc ctccctccac gggcccgggg 7200 gacaggtgtg cacgggccag ccaagggcac cttcgccacc ttcgagcggg cgaggaccgg 7260 gtggggacgg ggcggggacc cagctagcgg agctagagca gcctgcccgg ccacagcaca 7320 aaggaaagct agggcggggg aggagcgccg agggctgggg gccagcgtgc cccgcccacc 7380 gggggcttct cggggttgtc tccccccaca tctaggttag ggtgtgatac aaggaggttg 7440 attgccacat ggcaaccacg aagtgacttc tttatccctc acggatcgca gaggaagagg 7500 cagggctgag cccttctctc ccctgctgaa ggagcagatc gctaggacga ggcagatagg 7560 caggaagaga ctcaggagat ggacggagca gcttggctag gggggcagct gggag cgtgg 7620 agggagcctc cacagaggga tgccagatga gtagatctct gggaccctag cccctccact 7680 cagcagcagg gggctatcaa cctccgactg gggcagagtg agtgagtggc ttatctcccc 7740 ctccccagct gttaccctcc ctgagctgga ggagggtgtg agccacctgc cttggtctga 7800 agtctcaggg tgtcaggaaa accaaggggg ctccgatgcc ccattcaggg cacagccccc 7860 aagtgttctt ctcaaaaaaa aaaattgtat ttgaatcttt tcctctcctt aggctggaag 7920 taggctatcg ttgagaaaac cgaggccgag ttggggctga tctgagggag cctccagttg 7980 ggctggcctc actctcaccc tgggagggca accgggtagg ggggctggaa gatggaacaa 8040 aataaccagg cgtccaggcc atggtactgc agcactggcc ggtagccagg cccagaggga 8100 tggactcagg agacaagttc tcgtgtcttg caggtcagtt ataggagagg ccagcacagg 8160 gcaagcaggg cagtttccat cttattgtcc cagccctgcc cctggggggc tgagatcatg 8220 agaggcaatc tggacaagtg gcctggtgga ggagcctgtt tggggatgtg gccctgcctg 8280 aggagaaaca ctggccagtc tatggggtgt agacaacacc ttctatctct taacccctcg 8340 aaagctggac actggcaccc cagtgctggg gctactaatg acttctgccc cgcatcctgc 8400 tcagaacagg cccgggcctc catcaccagc agcatccatg tagccaaagg gaaggagtta 8460 ttcatccttg gtctgtgcgc cagccccatc tgggttgtgt gtgggacatg tgcagatgca 8520 cagatgcacc ctcgcttata tgtgtgctta cttccaaggg ccttaacggc atgttgtaaa 8580 agggtggtga gggtcccatc ctgaagaaga tggcgtggtg cttctcagct cttatcctac 8640 aaccagcagc tggctgagat ctcaaggctc aggacagacc cacattttag gagtccagca 8700 ggaggggtgt ctgggttcaa acgttgggtc atgagtattg gatgtcttcc ctgtaccttg 8760 actctgtgtg tgtgtgtgtg tgtgtgtgtg tgtgtgtgtg tatagtggag cttggggctg 8820 ctggccccta ggaggcagag ggtctcttaa atgcactctg tccttgtgct ataagcaggt 8880 cacaggggaa gtcagattgt gtggtgggaa aggccagcac caccctggtt atgatgtgat 8940 tcattgtgtg agaagttcct gctcttctca agtgctctga ggaagagggg gatgggcaga 9000 gtccatctgg ggtctcctgg gcctttccag ttccagctcc ctgagagcat gccggtcttg 9060 aaccttccac tgctagtatc tctgctgaag ccagactctc gccttgccct agtcaggtat 9120 ctctgcctta ggtggtagat aagtgttttt gtcaacatta caaatccctg tggtgctatt 9180 ttgtatagag gcagcagtgt gtgttagcac agtcagagtc cctgccttcc caagactgac 9240 aggacagcct gtggcagtgg ctatctccag ggcttgctgc aggtgaggtg tgcacatcta 9300 g agttctctt ctactttgcc ctgagtgttt ccgagctcct acctgccagt ctcagagtac 9360 ctgtgttatg ttagagaaac actgtccagt agggtctggg gctagtgcag atagatgtct 9420 ttcccggttc ctgctgagtg ctatagcact ctggggcgtg aagctcagcc tgaggctggc 9480 ccctggagtt ggtagcggta tcccgagtgg tcttcatgta caggaaaatg caatgcaact 9540 cttgggtaca aagtgttctt caaaactgaa tctgggagcc agcgtggccc aagtgtgggt 9600 gtgaatacac agtcattttg gagaggtggg ctaggcagag ctgagagggt ttgaatgaga 9660 cacagtctct gctgaggaag gattgtggcc ctagcaggga agagggacgg tttggaggtg 9720 ggtgggcatg gagaaatgat aaaaggggct tcagagtctt gagatgagca gcttgcccca 9780 gagagatgtc atggccagag cagtgtcttg gaaggagcca ggattcttgg tgctggctca 9840 gtgtttcatc cgtgactaca gagggagaca ccctatttct ttgagggctc tttgttcctg 9900 gccatggggt tagggcagcc tgacaggtgg tggggcagct gtgcggggaa agcagctgtc 9960 ggattggctg gggcagcagt tgcaccaagg gctcggagac atgcagccca cctgtgtctc 10020 tttgtggagc aaagcaggga gggatgcagg aaggggaggg gctgcaggga cttatttggt 10080 cgccttaaca ggggtcaata acccctgggg catggggtta gccctccctc gttccttcct 10140 ctcc tccttc tcctcctcct ccttctcctt ttttgctcct tttcttttct tccctcctcc 10200 tccatttttt attttttcca cttccctcct gtctcttgtc tgttctcttc cccctcttct 10260 tttcttcctc ctccctgctc ccccctttcc tccatctctg ctttttcatc ttctcgcaca 10320 ccccagcctc ctgacttaga tgggaaatgg gagatgagca aggttctgag cccttctgaa 10380 tctcagggac caagagtctc ttcttcatgt gtgtccagcg tggtgcctgc atcaggatgg 10440 gttgaggggt ggctcaaggg aagaataact acattacctt ccatgaaatg gatgcgttag 10500 tatgcctgcc aagaataaat acagagcaga aaaggctagg gaggaaggaa gtcattagtt 10560 tagctttctg gtcggcttca gtctatatgg tagtcctagg aggcttcagg gagggcagag 10620 gctgcctcag ggctagaaga ccagatctta gggtcagagt cggggagtgg tcacagcaac 10680 ctcatcccca tctcagagag ggagagattg agtctgagga tgtccccagg gtcctttgtg 10740 ttcgaggaga atgcagctct ggtgggctga tttctgtgcc aattgctggg ttgccatggt 10800 taccagggga tgcccagcat tgagcaccag attcttctcc ctcccaaaag ggggaggggc 10860 agcgggtacc aaggcacgca ccaggtagat tcctggctct ggccaatgag ggatgagaat 10920 catggtcgcc agggaacggt aaccaaggaa ccgttgcctt ggagtctcca tcgcacaccc 1 0980 gctccactga gttgtgggct gccagggcta gggccggacg ctgagctgac taggcagcga 11040 tttctccccc accgttccct gattgccatc tgacccagga ctcccggctc agcggaacga 11100 tgccatgcct agctttattg tcatgaccaa ccgacaacca cagctttccc cctcctcgtc 11160 ctgttctgcc ctgcctgtcc cctctggttt cactctccct ccctagtgcc ttccccaccc 11220 aaaattggct gcctaggccc tgacagttct gagccatcct ccctctcctc agggctctgt 11280 acacatacga ggatggctca gatgacctca agcttgcagc gtcaggaggt aagaattcca 11340 gccgttccct ctccttccca ccacccactc tgcttgtttt gttcccctct cgctttcctg 11400 cctccatcca gcagacccac gcttgtccac ggttgtcctt cctcttggtc attcccgcat 11460 tcctcagagt gtaggggggc ggggggggac tcctgcttcc tgtttgactt ctgggaaggc 11520 acagtgagga aaagcaactt gcccggcatc atacagctag ggaaggggcc tagtagggtg 11580 gtccagctca gtgctattct tggagctgtg agatttcctt aagcaaagct tcctgtacct 11640 tgctgggcac tagcacctgg aaagacacga gaggtgtgat ccccgagaaa tggagatgtg 11700 ttaacagacc tcttttgtcc catctcaagt caggcccagg aggattcaga ggcaaagata 11760 caatcctacc aaccccttcg cctcattaca cagaggtctg ggcctctagc tctg aggaat 11820 agaaatcccc tggcagggag gggttctagg cccttgcagg gtgacgggtg taggagatgg 11880 ggagtcccaa catctccctg gtttctccag aagggggctt gcaggagctt tccggccact 11940 tcgagaacca gaaagtgatg tatggtttct gcagcgtcaa ggactcccaa gctgccctgc 12000 caaaatatgt gctcatcaac tgggtatgtg gctgtgggat ggggctggga gctgaaggcc 12060 ccggagcggc cacattccac ccatagccac tgctggagga aaaagcctac actagttctt 12120 gggaagatag cctgggccca taggctgctt tgccttcagg ctgttgggcc agtgggctgt 12180 tgcccactcc tcccgtcctg agcccctctt cactgggcac attgcaggtt ggtgaggatg 12240 tgcctgatgc ccgaaaatgt gcttgcgcca gtcatgtggc caaggtggct gaattcttcc 12300 aggtatgttg ggaccaggtc tgaagtgggt acggttggtc acctagggcc aggtttgtat 12360 gctggggttg gtgatgggaa ttcggatacc acctaaacac tgctttttgt atggaagggt 12420 gttgatgtca ttgtgaatgc cagcagtgtg gaagacatcg atgctggtgc cattgggcag 12480 cggctctcca atggactggc acggctctcc agcccagtat tgcaccgcct gcgccttcgg 12540 gaggatgaaa atgctgaacc ggtggtcagt gtattcccgg gacaggctgg tctcgtgtca 12600 cctgtgtgac ctggtcccca gtcgggcctc cccaggccct cttagcc ctc attctgcaca 12660 tgtgagtact ggactgtgcc ttgggtctgc tcttgcgctc atgcccaggc acttgactct 12720 gcagggtacc acctaccaga agacggatgc agcagtggag atgaagcgga ttaaccgtga 12780 gcagttttgg gagcaggcca aggtgggcaa tgctgtagcc ttcaggccca ggtgtcccta 12840 cctgagcctg cggggtagct ctgacctctc tcggctgaaa gagcaaaccc ttttggtctc 12900 ctgcacacag ggctcatccc cctaattccc agaaggccac ctccccagcc tgggtgggag 12960 tagaattgga agggccgaag ctggcctggg ctcagggctt gtccttcctg ggtgtggggt 13020 gcagaaggag gaagagctgc ggaaggagga ggagcggaag aaggctctgg acgccaggct 13080 caggtttgaa caggaacgga tggagcagga gcggcaggag caggaagaac gtgagcggcg 13140 ctaccgggag cgggagcagc agattgagga gcacaggtaa gctcaggccc caggcgaagg 13200 gccaccccgc cccgccccgc cccgcccacc gcccccgccc accgccccag cctactgccc 13260 ccagtccacc ctgaaggggc ttccctgcac ttctccttcc aggaggaaac agcagagtct 13320 ggaagctgaa gaagccaaga ggaggttaaa ggagcagtct atctttgtaa gttcttttcc 13380 aggggtcttg tgcctggcca ggaacgtgag gctggtctgt gtgttctgcc ctctgagggc 13440 atgcgtgggc atccctatca tgagtgtgtc aggtccaaca cctatcttgt gggttcctgg 13500 tgtttgtgtg aaaaacaaac agacaaataa cagcagcagc aacaacaaaa catttttttt 13560 cgacgttttt ctccaaattg gaaaagaaaa tccaccttga ctcagaccct cgctgacctg 13620 gctcctccac tgcttccgcc ttgagacctg gggtccttgg ggtgtattag tcttgggctc 13680 tgcatttggg gctggggggc ttgagaatta ggtaggcttc ataatgaaag gactcagagc 13740 tcctgcctga atgtaggcca gtgttgttag ggactgaaca catagacagg ctctggggca 13800 gcctcctcct ccactgttgc tgtcccaatc tatattgtgc acagccattc acagcaagat 13860 cttttgaaaa agccacgttt tgtaggaatt tccaaggtgt gcgtgcacac agagaagtag 13920 attagtgtag tgacttctga atttgccctt catctttgtc agtcaccagc atcttaagca 13980 atggctgcag aggttttgct cttcttcaag agttaatcca ctgcttcaaa gtatctcata 14040 ggaaagtaac gatgtagcaa cattgtagcc agctgtgctc caagaaaggt cccaccggga 14100 gctgaaattt agcactctaa aatgatcccc caccaccacc caaaaatggg tttctctgta 14160 gccctggaac ttgctctata gaccaggctg tccttgaact cataaggatc aacctgacac 14220 tgcatcccaa gtgctgggtg ccaccactat ctagctaaaa tgatttaaat taggaatgcg 14280 taggcgctgt atgctctctc tcctgtcagt g agtttactt gtttactcag gtatatgagt 14340 gttctgcctg cctgtatgct tgtgcactgt gtgtctgcag cgcctgcaga attcagaaga 14400 tagtggctct cctagaactg gagttacaga tggttgtgag ccagcatgtg ggtgctggga 14460 accaaacctg ggtcctctac aagagcagca aggactccta accacagagc catctctcca 14520 gcccctgagt tcccttctgg ccttgggttc tcatgtgtca gaacacttag gcacagcaat 14580 gctttgaccc ctgagtcttc ctctccctcc tacctacagc tctaacttgt ttcagtgatg 14640 ttgacattat tcacagttaa cacctctctg ataagcataa tctcttcctt ttatggtgtg 14700 tgtgtgtgtg tgtgtgtgtg tgtgtgtgtg tgtgtgtgta tttgtggtat ggatagtgca 14760 tttacccatc atgcacttgt aaagaccaga ggtccgcttc tggtgtctcc ctctgtagct 14820 acgcactgta tttttttgag acaggctctc tctttgaacc tgcttactgt tcagctagtg 14880 agccccaggg ctctctgcct ctgctcaaat gttggtgata tagatctatg ttctctgccc 14940 agctcagtat gtgtcttgct gaggatccaa acttgagaag tacatgcttt acccacggag 15000 acaactctcc agcctatatc tgcccctttc tttgatggca gaaaaataag gtctaggtta 15060 tatgctcaaa ggtcaaactc tgagccagga gtggtggcac aagcctgtaa tcccaacacc 15120 aggaaagcag aggcagacag atca ctgtga cttcgaggcc agcctggtct aggctgtgag 15180 ctccagacca cccagctata taaagtaaga aatataagaa aaaggttctg ttcttcacct 15240 agctcttgcc ccttctgccc agtttgcccc atgaatgaca gaagatgcca actaagagac 15300 ttttgatata gagagttcca ttgaacaaac acataccagc cccatgctgg ccgttcacat 15360 agacttgggg cttcatccgg tgttgctcac tgagggtgag ggaagtgacc ccagaaacag 15420 gaaacacaat gctgtgcagt ctgtggtatc tgctgggagc tccctgaggg gacccagaaa 15480 aggaaatgac ctaaatgaac cagggacttc ttgctctgga gttagtgacc aagcagaggc 15540 cttaaaactg gagagactgg aagaggtgtg acgtgagctg gcctccgatg ccaaaggatg 15600 gggttgggaa acgggatgac gactctaggc tgtgactgga gcatgtagag gacattatgt 15660 ccacatctag cagaggatgt gggcgagttg gagccaggat agatggtggg gttggagacg 15720 tgaagaagac tagatcagac tgaggaaggg gaacggagcc aagcaagggg cctgcccacc 15780 tacagtcctg ggcttcagtc ttaaaacata agccaagaaa tggccaaatg ggaattaggg 15840 agtgggggac tgcagtgaac tgggctgtca ccactgtgta gggaaactag gggcttgagt 15900 gtgcagctca gaggcagagc gctgtccagc aagcctccac cccaactccc agggtgggac 15960 agagtcacac agccaga gag agagagagag agagagagga gagacagaga gagaatacaa 16020 actagatact ctgaggggct gtgagagatt ctgtctcaaa aaaaaaaaaa aaaaagaaaa 16080 aaaaaaaaac caaaacaaaa gccaggggag gtggtaaaac caaaagttca aggccagtct 16140 aacagtttaa tgagactctg tctcaaaata aaaccataaa aggtgtagtc actgctaaag 16200 cattcgcttg gcttctttga ggacctgggt acgatgcctc ggatgacaaa ataagaaacc 16260 agattctatt ttcagttcat catgaaactg aacttcttag tgatagcagc ttaatttatg 16320 tgggtttaaa accttgctat ttggggcctg aagagagggc tcagcagcag tgaagagcac 16380 cggtttctct tccagaagat ctgggttcaa tccccagcgc ccacatgtcc ggagccatct 16440 gtgtctccaa ttctagggat acaacgcctt cttttggcct ctgagggcac tgcatgcacc 16500 tggtgagcag acatccaggc aggcagaaca tccacacaca taaaaataat gtgttttgtt 16560 tttgttttct tttttttaat ggctttaaaa tatggcattg gaacaatacc tagtccatgg 16620 gccatattca gtccgtgctt ggtgtgactt aagctagcta agggaccagg aatattctga 16680 gtgatcccag atacctggtt agatgggacg tagatgaatc agcggctcaa ggcctggctg 16740 gccggtctgg ggatgaggtg gctgcagaca ttgaaagagg aggctgtgga cctgaattct 16800 gagaggaac a aagcgccacc tttgagattc tgcggggttg gaaagaacca ctcaggacat 16860 tgtccctttg gttttccagg gtgaccagcg ggatgaagag gaagagtccc agatgaagaa 16920 gtcggagtca gaggtggagg tgagagcggg gggagtggca agtgagacgg atgggcgggg 16980 cttggctcag actgctagcc ccgcccagct ctctaactct tcccttgtgc cctcaggagg 17040 cggctgccat cattgcccag cggcctgata acccacggga gttcttcaga cagcaggaac 17100 gagtggcatc ggcctctggt ggcagctgtg acgcgcctgc gcctgcaccc ttcaaccacc 17160 gaccaggtag tccttggccc cctccttgcc caatcagtac tcctcagact atggccctaa 17220 ttccctctgg gacttcttct tggaaaagaa gccttgccca aggaaggact ccttggagaa 17280 ggcaagaccc ccttcagtgt gcttccaccc ccagctttcc aggtggtggc acccccggag 17340 gggctgggtt ggggggcact gtcaatgtcc ggcattggct gagtatggct ggtactcaag 17400 gccagttggg aagatttggt gaagcttcag acacactgct ggtgttggga gtttccctgc 17460 agggttggac taggtctggc ccgaaggctt tggatgcatg gagggtggga tcttgctctg 17520 tcctgcagtg tgagcatcct ggagcagtgg ttctctcggc tcgcttgttt ggcttctcct 17580 tgtgtagctt ctgcctcccc ttggaagcac cctttcattt cttgtctgcc ggtgtgagca 17640 t ttgcatctt ctgtgtgctt ggtcatgttg ccggggttcc aggagcatgg cagactagac 17700 tcattcgtgc tgctacacag aggctggctg ttgtgtgccc atgtgtgtat ctggtgctgg 17760 ttggtgggag gcatcagatg gcttgaggct gtggcccagc ttggctgaat gtgcaccctg 17820 gctgaaggtg gcttggacat ctagggctgg ccctgggggt gggtgaggcc ttggggcagg 17880 ggtgggggag actgaccctc tcttgtcctc tgtctctttc tctgtctgtc tctctctcgg 17940 caggtcgtcc gtactgccct ttcataaagg catcggacag tgggccttcc tcctcctcct 18000 cttcctcctc ttcccctcca cggactccct ttccctatat cacctgccac cgcaccccaa 18060 acctctcttc ctccctccca tgtaggtagc agccccaggc ctcggcaggg tggggtaagg 18120 agggccccgc ttcccagcag ccccctcgcc ctttctcccc cagcctgact gccgactgct 18180 ggtgtttgtc cccagagcag gtactacctg cctcttgtcc ctggccctgt agggcactgg 18240 cccccagcgg cctggttcac agggtctgga gcagcaatct ggcccctggg gggcctgggg 18300 agctcacagc ccccacccac acttgagaca caaggtggat agtcctgatt ccacctggct 18360 cccacttcct cactcactag cctgggcatt tgctccgcag gcagccacct ggacagccac 18420 cggaggatgg cacccactcc tattcccacc cggagcccat ctgattccag cacagcctct 18480 acccccatcg ctgagcagat cgagagggcc ctggatgagg tcacatcctc gcagcctcca 18540 cctccacctc caccacctcc accaactcaa ggtaagagaa gggccccacc ggtacctatc 18600 cttccatcag tctctgggta gggaacccta ggttgccaca gtaacatagt tcaaagaact 18660 tggaccaatc ctcaccaaca ctacactgct aaggaagtgg ctatagcaga ggagcgccct 18720 ggtgttgctc ttagacaatg ccttctctaa ccctcagtgg ctcatcactg aggaatgtgg 18780 ccttcctcct ggtggtttaa aaaaagttgg tgagggctgg agagatggct ccgaggttaa 18840 gagcactggt tgctctccct gaggtcctga gttcaattcc cagcatccac atggtggctc 18900 acaaccacct ataatgagat ctggcaccct cttctgacct ggaggcgtac acgtaggcag 18960 gatgctgtat acataataaa aatgaatctt tcaaaacata aaaatgaaaa gagtgggtga 19020 aaggataggg aggtacagct gtagtagaag gtatctcaaa tttacccaga agtctcaaac 19080 tcgtcactct tcagtgggtg gacatggtac actggcccag ggcatctatc cttccccaag 19140 ggggctgact gtattgtgtc cttcatgtgc actgccacag aggcccagga gactacccca 19200 agcctggatg aagagctcag caaggaggcc aaagtaacag cagctcctga ggtctgggct 19260 ggctgtgcgg cagagccccc tcaggcacag gaacctcccc tgttgcaaag ca gccccctg 19320 gaggactcga tgtgcacaga atctccagag caggctgccc tggctgcccc tgcggagcct 19380 gctgcctctg tcacctcagt agctgatgtc catgcagctg acaccattga gaccaccact 19440 gccactactg acaccactat tgccaacaac gtcacccctg ccgctgccag cctcattgat 19500 ctatggcctg gcaacgggga agaggcctca acacttcagg ctgaacccag ggtgcccaca 19560 ccaccctcag gtgctgaggc ctccctggca gaggtgcccc tgctgaatga ggccgctcag 19620 gagccgctgc cgccggtagg cgaaggctgt gctaaccttc ttaattttga tgagctgcca 19680 gaacctccag ccaccttctg tgacccagag gaggaagtag gagaaacgct ggctgcctcc 19740 caggtcctaa ctatgccctc agctctagag gaggtagatc aggtgctgga gcaggagctg 19800 gagccagaac ctcacctgct gaccaatgga gagaccactc aaaaggaggg gacccaggtg 19860 ggacaggggg agcctggtgg gagggacagt attcagtgcg gagggggagg gcggattgct 19920 tgtagatcca tattgagaag cagctctatg ccacgcccct ttccatctct gccattcgtt 19980 cctggcaggc atctgtgttt ccccagcaac caaggattcc aagagtggtt gctggggaaa 20040 caggctcagg cctgatagat cagctgttgt cctgacagtt gggtcctggt gacctaagct 20100 tttattaaac taactaaata aacaaacaaa acaaaaccaa ataa caaacc ccagcaggcc 20160 agcgaaggat acttcagtca gtcacaggag gaagagttcg cccaatcaga agagccatgt 20220 gcaaaggttc cgcctcctgt attttacaac aagcctccag gtagtatccc cgggtgaatg 20280 atgggaattc tggaattctg ggatgggact ggagggctag gggtgtcact agagtcggtt 20340 ggcacagctc ttagaggttg gggggcgggt ggcaccactg atggatctac ccagccttct 20400 ctcttctctc aatatatcct agaaatcgac atcacttgct gggatgcaga cccagttcct 20460 gaagaggaag agggcttcga gggtggtgat tagtagcggc gactgccccc tggctgccct 20520 cgccaaggct gcctacctgc agtggcctct ggccagccgg cttgcagtgc cagcattagc 20580 agcagccccg cctggctccc actctggatt ccggcactgg ccggggacct gtctgcttcc 20640 ttacccacag ggcctgactt ttacagcttt tctc 20674 <210> 3 <211> 2124 <212> DNA <213> Rattus norvegicus <220> <221> CDS <222> (1) .. (2124) <400> 3 atg gcc ggc gtc agc ttc agc ggc cac cgc ctg gag ctg ctg gcg gcg 48 Met Ala Gly Val Ser Phe Ser Gly His Arg Leu Glu Leu Leu Ala Ala 1 5 10 15 tac gag gag gtg atc cgg gag gca gcc gac tgg gct ctg tac 96 Tyr Glu Glu Val Ile Arg Glu Glu Ser Ala Ala Asp Trp Ala Leu Tyr 20 25 30 act tac gaa gat ggc tca gat gac ctc aag ctt gca gca tca gga gaa 144 Thr Tyr Glu Asp Gly Ser Asp Asp Leu Lys Leu Ala Ala Ser Gly Glu 35 40 45 gga ggc ttg cag gag ctt tca ggc cac ttc gag aac cag aaa gtg atg 192 Gly Gly Leu Gln Glu Leu Ser Gly His Phe Glu Asn Gln Lys Val Met 50 55 60 tac ggt ttc tgc agc gtc aag gac tcc cag gct gcc ttg cca aaa tat 240 Tyr Gly Phe Cys Ser Val Lys Asp Ser Gln Ala Ala Leu Pro Lys Tyr 65 70 75 80 gtg ctc atc aac tgg gtt ggt gag gat gtg cct gat ccc aaa tgt 288 Val Leu Ile Asn Trp Val Gly Glu Asp Val Pro Asp Ala Arg Lys Cys 85 90 95 gct tgt gcc agc cat gtg gcc aag gtg gct gaa ttc ttc cag ggc gtt 336 Ala Cys Ala Ser His Val Ala Lys Val Ala Glu Phe Phe Gln Gly Val 100 105 110 gac gta att gtg aat gcc agc agt gtg gag gat atc gat gct ggc gcc 384 Asp Val Ile Val Asn Ala Ser Ser Val Glu Asp Ile Asp Ala Gly Ala 115 120 125 att ggg cag cgg ctc tcc aat gga ctg gct cgg ctc tct cca gtg 432 Ile Gly Gln Arg Leu Ser Asn Gly Leu Ala Arg Leu Ser Ser Pro Val 130 135 140 ctg cac cgc ctg cgc ctt cgg gag gat gag aat gct gag ccg gtg ggc 480 Leu His Arg Leu Arg Leu Arg Glu Asp Glu Ala Glu Pro Val Gly 145 150 155 160 acc acc tac cag aag acg gat gcg gcc gtg gag atg aag cgg att aac 528 Thr Thr Tyr Gln Lys Thr Asp Ala Ala Val Glu Met Lys Arg Ile Asn 165 170 175 cgt gag cag ttt tgg gag cag gcc aag aag gag gag gag ctg cgg aag 576 Arg Glu Gln Phe Trp Glu Gln Ala Lys Lys Glu Glu Glu Leu Arg Lys 180 185 190 gag gag gag cgg aag aag gct ttg gat gcc agg ctc agg ttt gag cag624 Glu Arg Lys Lys Ala Leu Asp Ala Arg Leu Arg Phe Glu Gln 195 200 205 gag cgg atg gag cag gag cgg cag gag cag gag gaa cgg gaa cgg cgc 672 Glu Arg Met Glu Gln Glu Arg Gln Glu Gln Glu Glu Arg Glu 210 215 220 tac cga gag cgg gag cag cag atc gag gag cac agg agg aaa cag cag 720 Tyr Arg Glu Arg Glu Gln Gln Ile Glu Glu His Arg Arg Lys Gln Gln 225 230 235 240 agt ctg gaa gct gaa gag gcc aag agg tta aag gat cag tct atc 768 Ser Leu Glu Ala Glu Glu Ala Lys Arg Arg Leu Lys Asp Gln Ser Ile 245 250 255 ttt ggt gat cag cga gat gaa gag gaa gag tcc cag atg aag aag tcg 816 Phe Gly Asp Glu Arg Asp Glu Glu Glu Ser Gln Met Lys Lys Ser 260 265 270 gaa tct gag gtg gag gag gca gct gcc atc att gcc cag cgg cct gat 864 Glu Ser Glu Val Glu Glu Ala Ala Ala Ile Ile Ala Gln Arg Pro Asp 275 280 285 aac cca cgg gag ttc ttc aga cag cag gaa cga gtg gct tca gcc tct 912 Asn Pro Arg Glu Phe Phe Arg Gln Gln Glu Arg Val Ala Ser Ala Ser 290 295 300 ggt ggc agc tgt gac gca ccc tcg ccc ttc aac cgtc cgt 960 Gly Gly Ser Cys Asp Ala Pro Ser Pro Phe Asn His Arg Pro Gly Arg 305 310 315 320 ccg tac tgc cct ttc ata aag gca tcg gac agt ggg cct tcc tcc tcc 1008 Pro Tyr Cys Pro Phe Ile Lys Ala Ser Asp Ser Gl y Pro Ser Ser Ser 325 330 335 tcc tct tcc tcc tct tcc cct cca cgg act ccc ttt ccc tat atc acc 1056 Ser Ser Ser Ser Ser Ser Pro Pro Arg Thr Pro Phe Pro Tyr Ile Thr 340 345 350 tgc cac cgc acc cca aac ctc tct tcc tcc ctc cca tgc agt cac ctg 1104 Cys His Arg Thr Pro Asn Leu Ser Ser Seru Leu Pro Cys Ser His Leu 355 360 365 gac agc cac cgg agg atg gcg ccc act ccc att ccc acc cgg agc cca 1152 Asp Ser His Arg Arg Met Ala Pro Thr Pro Ile Pro Thr Arg Ser Pro 370 375 380 tct gat tcc agc aca gcc tcc acc ccc atc acg gag cag atc gag agg 1200 Ser Asp Ser Ser Thr Thr Ala Ser Thr Pro Ile Thr Glu Gln Ile Glu Arg 385 390 395 400 gcc ctg gat gag gtc aca tcc tcg cag cct cca ccc cca cct cca cca 1248 Ala Leu Asp Glu Val Thr Ser Ser Gln Pro Pro Pro Pro Pro Pro Pro 405 410 415 ccc cca cca gct caa gag gcc cag gag tct gct ccc aga ctg gac ggt 1296 Pro Pro Pro Ala Gln Glu Ala Gln Glu Ser Ala Pro Arg Leu Asp Gly 420 425 430 gaa gag gtc tgc aag gag gcc aaa gta gca gca gct cct cag gtc tgg 1344 Glu Glu Val Cys Lys Glu Al a Lys Val Ala Ala Ala Pro Gln Val Trp 435 440 445 445 gct ggc tgt gca gag gag cct cct cgg gca cag gaa cct ccc ctg ttg 1392 Ala Gly Cys Ala Glu Glu Pro Pro Arg Ala Gln Glu Pro Pro Leu Leu 450 455 460 caa agc agc ccc acg gag gac ttg atg tgc aca gaa tct cct gag cag 1440 Gln Ser Ser Pro Thr Glu Asp Leu Met Cys Thr Glu Ser Pro Glu Gln 465 470 470 475 480 gct gtc ctg gcg gcc tct cca gag cct gat gcc tct tca gta 1488 Ala Val Leu Ala Ala Ser Pro Glu Pro Asp Ala Ser Val Thr Ser Val 485 490 495 gct gat gct cat gca gct gac acc atc gag acc acc act gcc act act 1536 Ala Asp Ala His Ala Ala Asp Thr Ile Glu Thr Thr Thr Ala Thr Thr 500 505 510 gcc acc act att gcc gac aac gtc acc cct gcc gcc gcc agc ctc att 1584 Ala Thr Thr Ile Ala Asp Asn Val Thr Pro Ala Ala Ala Ser Leu Ile 515 520 525 gat ctg tgg cct ggt aac ggg gaa gag gcc tca acg cct cag gct gaa 1632 Asp Leu Trp Pro Gly Asn Gly Glu Glu Ala Ser Thr Pro Gln Ala Glu 530 535 540 ccc agg gtg ccc acc cca ccc tca ggt gct gag gcc tcc ctg gca gag 1680 Pro Arg Val Pro Thr Pro Pro Ser Gly Ala Glu Ala Ser Leu Ala Glu 545 550 555 560 gtg cct ctg ttg aat gag gca gct cag gag ccg ctg cca cca gtg ggc 1728 Val Pro Leu Leu Asn Glu Ala Ala Gln Glu Pro Leu Pro Pro Val Gly 565 570 575 gaa ggc tgt gcc aac ctt ctt aat ttc gat gag ctg cca gaa cct cca 1776 Glu Gly Cys Ala Asn Leu Leu Asn Phe Asp Glu Leu Pro Glu Pro Pro 580 585 590 gcc acc ttc tgt gac cca gag gag gaa gga gag ccg ctg gct 1824 Ala Thr Phe Cys Asp Pro Glu Glu Glu Ala Glu Gly Glu Pro Leu Ala 595 600 605 gcc tcc cag gtc cta act atg ccc tca gcc cta gag gag gta gat cag 1872 Ala Ser Gln Val Leu Thr Met Pro Ser Ala Leu Glu Glu Val Asp Gln 610 615 620 gtg ttg gag cag gag ttg gag cca gaa cct cat ctg ctg acc aat gga 1920 Val Leu Glu Gln Glu Leu Glu Pro Glu Pro His Leu Leu Thr Asn Gly 625 630 630 635 640 gag acc act cag aag gag ggg acc cag cag gcc agc gaa gga tac ttc 1968 Glu Thr Thr Gln Lys Glu Gly Thr Gln Gln Ala Ser Glu Gly Tyr Phe 645 650 655 agt cag tca cag gag gaa gaa ttc gcc caa tca gaa gag ca tgt gca 2016 Ser Gln Ser Gln Glu Glu Glu Phe Ala Gln Ser Glu Glu Pro Cys Ala 660 665 670 aag gct cca cct cct gta ttc tac aac aag cct cca gaa atc gac atc 2064 Lys Ala Pro Pro Pro Val Phe Tyr Asn Lys Pro Pro Glu Ile Asp Ile 675 680 685 acc tgc tgg gat gca gac cca gtt cct gaa gag gaa gag ggc ttc gag 2112 Thr Cys Trp Asp Ala Asp Pro Val Pro Glu Glu Glu Glu Gly Phe Glu 690 695 700 ggt ggt gat tag 2124 Gly Gly Asp 705 <210> 4 <211> 707 <212> PRT <213> Rattus norvegicus <400> 4 Met Ala Gly Val Ser Phe Ser Gly His Arg Leu Glu Leu Leu Ala Ala 1 5 10 15 Tyr Glu Glu Val Ile Arg Glu Glu Ser Ala Ala Asp Trp Arp Leu Tyr 20 25 30 Thr Tyr Glu Asp Gly Ser Asp Asp Leu Lys Leu Ala Ala Ser Gly Glu 35 40 45 Gly Gly Leu Gln Glu Leu Ser Gly His Phe Glu Asn Gln Lys Val Met 50 55 60 Tyr Gly Phe Cys Ser Val Lys Asp Ser Gln Ala Ala Leu Pro Lys Tyr 65 70 75 80 Val Leu Ile Asn Trp Val Gly Glu Asp Val Pro Asp Ala Arg Lys Cys 85 90 95 Ala Cys Ala Ser His Val Ala Lys Val Ala Glu Phe Phe Gln Gly Val 100 105 110 Asp Val Ile Val Asn Ala Ser Ser Val Glu Asp Ile Asp Ala Gly Ala 115 120 125 Ile Gly Gln Arg Leu Ser Asn Gly Leu Ala Arg Leu Ser Ser Pro Val 130 135 140 Leu His Arg Leu Arg Leu Arg Glu Asp Glu Asn Ala Glu Pro Val Gly 145 150 155 160 Thr Thr Tyr Gln Lys Thr Asp Ala Ala Val Glu Met Lys Arg Ile Asn 165 170 175 Arg Glu Gln Phe Trp Glu Gln Ala Lys Lys Glu Glu Glu Leu Arg Lys 180 185 190 Glu Glu Glu Glu Arg Lys Lys Ala Leu Asp Ala Arg Leu Arg Phe Glu Gln 195 200 205 Glu Arg Met Gl u Gln Glu Arg Gln Glu Gln Glu Glu Arg Glu Arg Arg 210 215 220 Tyr Arg Glu Arg Glu Gln Gln Ile Glu Glu His Arg Arg Lys Gln Gln 225 230 235 240 Ser Leu Glu Ala Glu Glu Ala Lys Arg Arg Leu Lys Asp Gln Ser Ile 245 250 255 Phe Gly Asp Gln Arg Asp Glu Glu Glu Glu Ser Gln Met Lys Lys Ser 260 265 270 Glu Ser Glu Val Glu Glu Ala Ala Ala Ila Ile Ile Ala Gln Arg Pro Asp 275 280 285 285 Asn Pro Arg Glu Phe Phe Arg Gln Gln Glu Arg Val Ala Ser Ala Ser 290 295 300 Gly Gly Ser Cys Asp Ala Pro Ser Pro Phe Asn His Arg Pro Gly Arg 305 310 315 320 Pro Tyr Cys Pro Phe Ile Lys Ala Ser Asp Ser Gly Pro Ser Ser Ser 325 330 335 Ser Ser Ser Ser Ser Ser Pro Pro Arg Thr Pro Phe Pro Tyr Ile Thr 340 345 350 Cys His Arg Thr Pro Asn Leu Ser Ser Ser Leu Pro Cys Ser His Leu 355 360 365 Asp Ser His Arg Arg Met Ala Pro Thr Pro Ile Pro Thr Arg Ser Pro 370 375 380 Ser Asp Ser Ser Thr Ala Ser Thr Pro Ile Thr Glu Gln Ile Glu Arg 385 390 395 400 Ala Leu Asp Glu Val Thr Ser Ser Gln Pro Pro Pro Pro Pro Pro 405 410 415 Pro Pro Pro A la Gln Glu Ala Gln Glu Ser Ala Pro Arg Leu Asp Gly 420 425 430 Glu Glu Val Cys Lys Glu Ala Lys Val Ala Ala Ala Pro Gln Val Trp 435 440 445 Ala Gly Cys Ala Glu Glu Pro Pro Arg Ala Gln Glu Pro Pro Leu Leu 450 455 460 Gln Ser Ser Pro Thr Glu Asp Leu Met Cys Thr Glu Ser Pro Glu Gln 465 470 475 480 Ala Val Leu Ala Ala Ser Pro Glu Pro Asp Ala Ser Val Thr Ser Val 485 490 495 Ala Asp Ala His Ala Ala Asp Thr Ile Glu Thr Thr Thr Ala Thr Thr 500 505 510 Ala Thr Thr Ile Ala Asp Asn Val Thr Pro Ala Ala Ala Ser Leu Ile 515 520 525 Asp Leu Trp Pro Gly Asn Gly Glu Glu Ala Ser Thr Pro Gln Ala Glu 530 535 540 Pro Arg Val Pro Thr Pro Pro Ser Gly Ala Glu Ala Ser Leu Ala Glu 545 550 555 560 Val Pro Leu Leu Asn Glu Ala Ala Gln Glu Pro Leu Pro Pro Val Gly 565 570 575 575 Glu Gly Cys Ala Asn Leu Leu Asn Phe Asp Glu Leu Pro Glu Pro Pro 580 585 590 Ala Thr Phe Cys Asp Pro Glu Glu Glu Ala Glu Gly Glu Pro Leu Ala 595 600 605 Ala Ser Gln Val Leu Thr Met Pro Ser Ala Leu Glu Glu Val Asp Gln 610 615 620 Val Leu Glu Gln Glu Leu Glu Pro Glu Pro His Leu Leu Thr Asn Gly 625 630 635 640 Glu Thr Thr Gln Lys Glu Gly Thr Gln Gln Ala Ser Glu Gly Tyr Phe 645 650 655 Ser Gln Ser Gln Glu Glu Glu Phe Ala Gln Ser Glu Glu Pro Cys Ala 660 665 670 Lys Ala Pro Pro Pro Val Phe Tyr Asn Lys Pro Pro Glu Ile Asp Ile 675 680 685 Thr Cys Trp Asp Ala Asp Pro Val Pro Glu Glu Glu Glu Gly Phe Glu 690 695 700 Gly Gly Asp 705 <210> 5 <211> 24 <212> DNA <213> Rattus norvegicus <400> 5 aggcatcgga cagtgggcct tcct 24 <210> 6 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Antisense oligonucleotide <400> 6 aggaaggccc actgtccgat gcct 24 <210> 7 <211> 9 <212> PRT <213> Rattus norvegicus <400> 7 Lys Ala Ser Asp Ser Gly Pro Ser Ser 1 5 <210> 8 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Antisense oligonucleotide <400> 8 ttatgaaagg gcagtacgga cgac 24 <210> 9 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Antisense oligonucleotide <400> 9 tatagggaaa gggagtccgt ggag 24 <210> 10 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Antisense oligonucleotide <400> 10 ggtttggggt gcggtggcag gtga 24

[Brief description of the drawings]

FIG. 1 is a diagram showing the exon-intron configuration of the mouse drebrin A genomic gene.

FIG. 2 is a view showing a chromosome map of a mouse drebrin A genomic gene (Dbn1) on chromosome 13.

FIG. 3: Antisense strand 5′-AGGAAGGCCCA
It is a figure which shows the result of the western blot when administering CTGTCCGATGCCT-3 '.

FIG. 4: Antisense strand 5′-AGGAAGGCCCA
It is a figure which shows the result of the immunostaining when administering CTGTCCGATGCCT-3 '.

FIG. 5: Antisense strand 5′-AGGAAGGCCCA
It is a figure which shows the measurement result of spine length at the time of administering CTGTCCGATGCCT-3 '.

FIG. 6: Antisense strand 5′-AGGAAGGCCCA
It is a figure which shows the measurement result of spine density at the time of administering CTGTCCGATGCCT-3 '.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) A61P 25/00 A61P 43/00 105 4H045 43/00 105 C07K 16/18 C07K 16/18 C12P 21/08 / / C12P 21/08 C12N 15/00 ZNAA F-term (reference) 4B024 AA01 CA01 HA17 4B064 AG27 CA19 CC24 4C084 AA13 MA01 NA14 ZA012 ZB212 4C085 AA13 AA14 CC32 EE01 GG01 4C086 AA01 AA02 AA03 CA14 A01 MA04 NA04

Claims (8)

[Claims] 1. An antisense oligonucleotide comprising the nucleotide sequence of SEQ ID NO: 6. 2. An antisense oligonucleotide which hybridizes with the nucleotide sequence of SEQ ID NO: 5 under stringent conditions and has an effect of suppressing drebrin A expression. 3. A derivative of the antisense oligonucleotide according to claim 1 or 2. 4. A vector carrying the antisense oligonucleotide according to claim 1 or 2 and / or a derivative of the antisense oligonucleotide according to claim 3. 5. An antibody that specifically recognizes a peptide consisting of the amino acid sequence shown in SEQ ID NO: 7. 6. The antibody according to claim 5, which is a monoclonal antibody. 7. A therapeutic agent for a disease caused by overexpression and hyperfunction of drebrin A, comprising an antisense oligonucleotide according to claim 1 or 2 and / or a derivative of the antisense oligonucleotide according to claim 3 as an active ingredient. . 8. An overexpression of drebrin A comprising the antisense oligonucleotide according to claim 1 or 2 and / or a derivative of the antisense oligonucleotide according to claim 3 and a pharmaceutically acceptable reagent for introduction into a cell. Drug for treating diseases caused by hyperactivity.