质粒名称: | pCS2-Cas9 |
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出品公司: | 爱迪基因 |
目录编号: | 47322 |
存储实验室: | Alex Schier |
载体骨架基本信息 |
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载体名称: | pCS2 |
载体出品公司: | -- |
载体抗性: | 氨苄青霉素 |
空载体大小: | 4095 bp |
完整质粒大小: | -- |
载体修饰: | -- |
载体类型: | 哺乳动物表达,CRISPR;爪蟾和斑马鱼的表达 |
筛选标记: | -- |
高拷贝/低拷贝: | 高拷贝 |
生长条件: | 37 ℃ |
插入基因信息 |
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插入基因名称: | Cas9 |
物种来源: | -- |
插入大小: | 4100 bp |
基因突变: | -- |
GenBank编号 | -- |
启动子 | SP6 |
标签/融合蛋白: | -- |
克隆方法: | 限制性内切酶 |
5' 克隆位点: | ClaI(未破坏) |
3' 克隆位点: | XbaI(未破坏) |
5' 测序引物 | SP6 |
3' 测序引物 | M13R |
文献引用方法 |
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材料和方法部分: | pCS2-Cas9 was a gift from Alex Schier (Addgene plasmid # 47322 ; http://n2t.net/addgene:47322 ; RRID:Addgene_47322) |
参考文献部分: | Efficient mutagenesis by Cas9 protein-mediated oligonucleotide insertion and large-scale assessment of single-guide RNAs. Gagnon JA, Valen E, Thyme SB, Huang P, Ahkmetova L, Pauli A, Montague TG, Zimmerman S, Richter C, Schier AF. PLoS One. 2014 May 29;9(5):e98186. doi: 10.1371/journal.pone.0098186. eCollection 2014. PONE-D-14-14210 [pii] PubMed 24873830 |
买家导航
pCS2Cas9载体基本信息
pCS2Cas9载体图谱质粒图谱和多克隆位点信息
pCS2Cas9载体简介
pCS2Cas9载体序列
LOCUS pCS2-Cas9 8215 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION For in vivo expression of Cas9 or generation of Cas9 mRNA. ACCESSION . VERSION . KEYWORDS pCS2-Cas9 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8215) AUTHORS Gagnon JA, Valen E, Thyme SB, Huang P, Ahkmetova L, Pauli A, Montague TG, Zimmerman S, Richter C, Schier AF TITLE Efficient mutagenesis by Cas9 protein-mediated oligonucleotide insertion and large-scale assessment of single-guide RNAs. JOURNAL PLoS One. 2014 May 29;9(5):e98186. doi: 10.1371/journal.pone.0098186. eCollection 2014. PUBMED 24873830 REFERENCE 2 (bases 1 to 8215) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8215 /organism="synthetic DNA construct" /mol_type="other DNA" protein_bind 107..128 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 143..173 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 181..197 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." primer_bind 186..208 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 205..221 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 205..221 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind 240..260 /label=T3 /note="T3 promoter, forward primer" promoter 242..260 /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" polyA_signal 373..507 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(410..429) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 464..483 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" promoter 512..529 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS complement(540..560) /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS complement(573..4676) /codon_start=1 /product="Cas9 (Csn1) endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system" /label=Cas9 /note="generates RNA-guided double strand breaks in DNA" /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL DATLIHQSITGLYETRIDLSQLGGD" regulatory 4673..4682 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" promoter complement(4721..4739) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(4722..4739) /label=SP6 /note="SP6 promoter, forward primer" promoter complement(4774..5751) /label=CMV IE94 promoter /note="enhancer/promoter region of simian cytomegalovirus major immediate early transcription unit IE94" rep_origin 5829..6284 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(5916..5935) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 6126..6147 /label=F1ori-F /note="F1 origin, forward primer" promoter 6310..6414 /gene="bla" /label=AmpR promoter CDS 6415..7275 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind complement(6633..6652) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 7446..8034 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 7935..7954 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 8188..8205 /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc 61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc 121 tcactcatta ggcaccccag gctttacact ttatgctccc ggctcgtatg ttgtgtggaa 181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg 241 caattaaccc tcactaaagg gaacaaaagc tgggtaccgg gcccaatgca ttggcgccgc 301 ggccgcgaat taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca 361 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc 421 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc 481 atcaatgtat cttatcatgt ctggatctac gtaatacgac tcactatagt tctagattac 541 accttcctct tcttcttggg gtcagccctg ctgtctccac cgagctgaga gaggtcgatt 601 cttgtttcat agagccccgt aattgactga tgaatcagtg tggcgtccag gacctccttt 661 gtagaggtgt accgctttct gtctatggtg gtgtcgaagt acttgaaggc tgcaggcgcg 721 cccaagttgg tcagagtaaa caagtggata atgttttctg cctgctccct gatgggctta 781 tccctgtgct tattgtaagc agaaagcacc ttatcgaggt tagcgtcggc gaggatcact 841 cttttggaga attcgcttat ttgctcgatg atctcatcaa ggtagtgttt gtgttgttcc 901 acgaacagct gcttctgctc attatcttcg ggagaccctt tgagcttttc atagtggctg 961 gccagataca agaaattaac gtatttagag ggcagtgcca gctcgttacc tttctgcagc 1021 tcgcccgcac tagcgagcat tcgtttccgg ccgttttcaa gctcaaagag agagtacttg 1081 ggaagcttaa tgatgaggtc ttttttgacc tctttatatc ctttcgcctc gagaaagtcg 1141 atggggtttt tttcgaagct tgatcgctcc atgattgtga tgcccagcag ttccttgacg 1201 cttttgagtt ttttagactt ccctttctcc actttggcca caaccagtac actgtaagcg 1261 actgtaggag aatcgaatcc gccgtatttc ttggggtccc aatctttttt gcgtgcgatc 1321 agcttgtcgc tgttcctttt cgggaggata ctttccttgg agaagcctcc ggtctgtact 1381 tcggtctttt taacgatgtt cacctgcggc atggacagga ccttccggac tgtcgcgaaa 1441 tccctaccct tgtcccacac gatttctcct gtttctccgt ttgtttcgat aagtggtcgc 1501 ttccgaatct ctccattggc cagtgtaatc tcggtcttga aaaaattcat aatattgctg 1561 taaaagaagt acttagcggt ggccttgcct atttcctgct cagactttgc gatcattttc 1621 ctaacatcgt acactttata gtctccgtaa acaaattcag attcaagctt gggatatttt 1681 ttgataagtg cagtgcctac cactgcattc aggtaggcat catgcgcatg gtggtaattg 1741 ttgatctctc tcaccttata aaactgaaag tcctttctga aatctgagac cagcttagac 1801 ttcagagtaa taactttcac ctctcgaatc agtttgtcat tttcatcgta cttggtgttc 1861 atgcgtgaat cgagaatttg ggccacgtgc ttggtgatct ggcgtgtctc aacaagctgc 1921 cttttgatga agccggcttt atccaactca gacaggccac ctcgttcagc cttagtcaga 1981 ttatcgaact tccgttgtgt gatcagtttg gcgttcagca gctgccgcca ataatttttc 2041 attttcttga caacttcttc tgaggggacg ttatcactct tccctctatt tttatcggat 2101 cttgtcaaca ctttattatc aatagaatca tctttgagaa aagactgggg cacgatatga 2161 tccacgtcgt agtcggagag ccgattgatg tccagttcct gatccacgta catgtccctg 2221 ccgttctgca ggtagtacag gtagagcttc tcattctgaa gctgggtgtt ttcaactggg 2281 tgttccttaa ggatttggga ccccagttct tttataccct cttcaatcct cttcatcctt 2341 tccctactgt tcttctgtcc cttctgggta gtttggttct ctcgggccat ctcgataacg 2401 atattctcgg gcttatgcct tcccattact ttgacgagtt catccacgac cttaacggtc 2461 tgcagtattc cctttttgat agctgggcta cctgcaagat tagcgatgtg ctcgtgaaga 2521 ctgtccccct ggccagaaac ttgtgctttc tggatgtcct ccttaaaggt gagagagtca 2581 tcatggatca actgcatgaa gttccggttg gcaaatccat cggacttaag aaaatccagg 2641 attgtctttc cactctgctt gtctcggatc ccattgatca gttttcttga cagccgcccc 2701 catcctgtat atcggcgcct cttgagctgt ttcatgactt tgtcgtcgaa gagatgagcg 2761 taagttttca agcgttcttc aatcatctcc ctatcttcaa acaacgtaag ggtgaggaca 2821 atgtcctcaa gaatgtcctc gttctcctca ttgtccagga agtccttgtc tttaatgatt 2881 ttcaggagat cgtgatacgt tcccagggat gcgttgaagc gatcctccac tccgctgatt 2941 tcaacagagt cgaaacattc aatctttttg aaatagtctt ctttgagctg tttcacggta 3001 actttccggt tcgtcttgaa gaggaggtcc acgatagctt tcttctgctc tccagacagg 3061 aatgctggct ttctcatccc ttctgtgacg tatttgacct tggtgagctc gttataaact 3121 gtgaagtact cgtacagcag agagtgttta ggaagcacct tttcgttagg cagattttta 3181 tcaaagttag tcatcctttc gatgaaggac tgggcagagg cccccttatc cacgacttcc 3241 tcgaagttcc agggagtgat ggtctcttct gatttgcgag tcatccacgc gaatctggaa 3301 tttccccggg cgagggggcc tacatagtag ggtatccgaa atgtgaggat tttctcaatc 3361 ttttccctgt tatctttcaa aaaggggtag aaatcctctt gccgcctgag gatagcgtgc 3421 agttcgccca ggtgaatctg gtgggggatg cttccattgt cgaaagtgcg ctgtttgcgc 3481 aacagatctt ctctgttaag ctttaccagc agctcctcgg tgccgtccat tttttccaag 3541 atgggcttaa taaatttgta aaattcctcc tggcttgctc cgccgtcaat gtatccggcg 3601 tagccatttt tagactgatc gaagaaaatt tccttgtact tctcaggcag ttgctgtctg 3661 acaagggcct tcagcaaagt caagtcttgg tggtgctcat catagcgctt gatcatacta 3721 gcgctcagcg gagctttggt gatctccgtg ttcactcgca gaatatcact cagcagaatg 3781 gcgtctgaca ggttctttgc cgccaaaaaa aggtctgcgt actggtcgcc gatctgggcc 3841 agcagattgt cgagatcatc atcgtaggtg tctttgctca gttgaagctt ggcatcttcg 3901 gccaggtcga agttagattt aaagttgggg gtcagcccga gtgacagggc gataagatta 3961 ccaaacaggc cgttcttctt ctccccaggg agctgtgcga tgaggttttc gagccgccgg 4021 gatttggaca gcctagcgct caggattgct ttggcgtcaa ctccggatgc gttgatcggg 4081 ttctcttcga aaagctgatt gtaagtctga accagttgga taaagagttt gtcgacatcg 4141 ctgttgtctg ggttcaggtc cccctcgatg aggaagtgtc cccgaaattt gatcatatgc 4201 gccagcgcga gatagatcaa ccgcaagtca gccttatcag tactgtctac aagcttcttc 4261 ctcagatgat atatggttgg gtacttttca tggtacgcca cctcgtccac gatattgcca 4321 aagattgggt ggcgctcgtg ctttttatcc tcctccacca aaaaggactc ctccagccta 4381 tggaagaaag agtcatccac cttagccatc tcattactaa agatctcctg caggtagcag 4441 atccgattct ttctgcgggt atatctgcgc cgtgctgttc ttttgagccg cgtggcttcg 4501 gccgtctccc cggagtcgaa caggagggcg ccaatgaggt tcttctttat gctgtggcga 4561 tcggtattgc ccagaacttt gaattttttg ctcggcacct tgtactcgtc cgtaatgacg 4621 gcccagccga cgctgtttgt gccgatatcg agcccaatgg agtacttctt gtccatggtg 4681 gcatcgatgg gatcctgcaa aaagaacaag tagcttgtat tctatagtgt cacctaaatc 4741 aagcttgctc cgacgtcccc aggcagaatg gcggttccct aaacgagcat tgcttatata 4801 gacctcccat taggcacgcc taccgcccat ttacgtcaat ggaacgccca tttgcgtcat 4861 tgcccctccc cattgacgtc aatggggatg tacttggcag ccatcgcggg ccatttaccg 4921 ccattgacgt caatgggagt actgccaatg taccctggcg tacttccaat agtaatgtac 4981 ttgccaagtt actattaata gatattgatg tactgccaag tgggccattt accgtcattg 5041 acgtcaatag ggggcgtgag aacggatatg aatgggcaat gagccatccc attgacgtca 5101 atggtgggtg gtcctattga cgtcaatggg cattgagcca ggcgggccat ttaccgtaat 5161 tgacgtcaat gggggaggcg ccatatacgt caataggacc gcccatatga cgtcaatagg 5221 aaagaccata tatagagacc attgacgtca atgggggagt ggctatgggc ggtattagga 5281 agccccatat atggtatatg ggaccgccca ttgggagggg ctatctacgt caataggaaa 5341 acccatatat ggaatactat atggcatagg gccaatacat agtattgaac ctggccaata 5401 gccatattgg catagggcca tattggatat tgcctatata ttgatcctgg catatagcca 5461 atatggccgc cattattggc accatgccaa ttcaatatgg cggacttggc acagggccaa 5521 gacccctcct caaacttggc accgtgccaa gtagacccct ccccagttgg cacagtgcca 5581 ggtccgccat attgaattgg catggtgcca agtccgccat attgaattgg ctggggtcca 5641 gatccaccat attgaattgg cacgagtcca tatacgccat attgaattgg cacaggtcca 5701 gatccaccat attgaattgg catgagtcca tatacgccat attgaattgg cggtcgactg 5761 gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg 5821 cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag 5881 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt 5941 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt 6001 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg 6061 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt 6121 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt 6181 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca 6241 aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg 6301 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg 6361 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt 6421 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt 6481 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg 6541 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa 6601 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt 6661 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag 6721 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt 6781 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga 6841 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt 6901 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta 6961 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg 7021 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc 7081 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt 7141 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg 7201 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg 7261 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa 7321 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa 7381 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga 7441 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg 7501 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact 7561 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac 7621 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg 7681 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg 7741 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga 7801 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc 7861 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg 7921 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc 7981 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc 8041 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt 8101 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc 8161 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaag //