质粒名称: | pSAG1::CAS9-U6::sgUPRT |
---|---|
出品公司: | 爱迪基因 |
目录编号: | 54467 |
存储实验室: | David Sibley |
载体骨架基本信息 |
|
载体名称: | pBlueScript |
载体出品公司: | Agilent |
载体抗性: | 氨苄青霉素 |
空载体大小: | -- |
完整质粒大小: | -- |
载体修饰: | -- |
载体类型: | CRISPR,弓形虫表达 |
筛选标记: | -- |
高拷贝/低拷贝: | 高拷贝 |
生长条件: | 37 ℃ |
插入基因信息 |
|
插入基因名称: | SAG1 5' UTR |
物种来源: | Toxoplasma gondii |
插入大小: | -- |
基因突变: | -- |
GenBank编号 | -- |
启动子 | -- |
标签/融合蛋白: | -- |
克隆方法: | -- |
5' 克隆位点: | -- |
3' 克隆位点: | -- |
5' 测序引物 | -- |
3' 测序引物 | -- |
文献引用方法 |
|
材料和方法部分: | pSAG1::CAS9-U6::sgUPRT was a gift from David Sibley (Addgene plasmid # 54467 ; http://n2t.net/addgene:54467 ; RRID:Addgene_54467) |
参考文献部分: | Efficient Gene Disruption in Diverse Strains of Toxoplasma gondii Using CRISPR/CAS9. Shen B, Brown KM, Lee TD, Sibley LD. MBio. 2014 May 13;5(3). pii: e01114-14. doi: 10.1128/mBio.01114-14. 10.1128/mBio.01114-14 PubMed 24825012 |
买家导航
载体基本信息
载体图谱质粒图谱和多克隆位点信息
载体简介
载体序列
LOCUS pSAG1::CAS9-U6::sgUPRT 9674 bp ds-DNA circular SYN 06-AUG-2018 DEFINITION Encodes CAS9 nuclease (GFP fusion) under control of the Toxoplasma gondii SAG1 promotor. Vector also provides U6 controlled expression of a single guide RNA for disruption of genes by CRISPR.. ACCESSION . VERSION . KEYWORDS pSAG1::CAS9-U6::sgUPRT SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 9674) AUTHORS Shen B, Brown KM, Lee TD, Sibley LD TITLE Efficient Gene Disruption in Diverse Strains of Toxoplasma gondii Using CRISPR/CAS9. JOURNAL MBio. 2014 May 13;5(3). pii: e01114-14. doi: 10.1128/mBio.01114-14. PUBMED 24825012 REFERENCE 2 (bases 1 to 9674) AUTHORS . TITLE Direct Submission JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..9674 /organism="synthetic DNA construct" /mol_type="other DNA" misc_RNA 559..633 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" promoter complement(658..676) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(697..713) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(697..713) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(710..732) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 721..737 /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)." promoter complement(745..775) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 790..811 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(928..945) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(1099..1687) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1179..1198) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(1858..2718) /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 2481..2500 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(2719..2823) /gene="bla" /label=AmpR promoter rep_origin complement(2849..3304) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2986..3007) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3198..3217 /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3431..3453 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 3445..3462 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 3446..3462 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 3472..3491 /label=T7 /note="T7 promoter, forward primer" promoter 3472..3490 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind 3515..3531 /label=pBluescriptKS /note="For pBluescript vector" primer_bind 3516..3532 /label=KS primer /note="common sequencing primer, one of multiple similar variants" CDS 4358..8461 /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" CDS 8465..8491 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" CDS 8510..8530 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 8537..8557 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 8597..9313 /codon_start=1 /product="enhanced GFP with monomerizing A206K mutation (Zacharias et al., 2002)" /label=mEGFP /note="mammalian codon-optimized" /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDDG TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE FVTAAGITLGMDELYK" primer_bind complement(8639..8660) /label=EGFP-N /note="EGFP, reverse primer" primer_bind 9247..9268 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(9635..9651) /label=SK primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(9635..9651) /label=pBluescriptSK /note="For pBluescript vector" ORIGIN 1 caagtaagca gaagcacgct gtatttccgg gagggtgcga cgagacaaag tgcgcgagtt 61 gaaatcgtcg tggggacgat tgcaccgcgg ccacatgttg gagacactga ggacacacgg 121 gaaacgcgaa agatttcaaa ttaacgtacc caagcgcgaa agcttgcgca gcatacactc 181 gaagcgaaca tcccgaacca tcgagaggcg gagagcgata agtctttcac gctgcgaagt 241 gttgcgacgg ctgcgccgct gcactgtgaa ttgggcgcca atattgcatc ctaggcctga 301 cgcgcctcct gcagaacgcg agacactggg atatgtagag ccaaggggga aaccttcgaa 361 ctctcgaatg tcttctctga caagaatcat atttccatca gttctgtcag attttcaaat 421 ggcgacctgc agaggcctgc ttcctccctg tgcgctcttc gaaggggctt tctgtcgcgc 481 agggtcacct cgtccccgaa gggggtgttt gccttctggt aaatggggat gtcaagttgg 541 cgtctcgatt gtgagagcgt tttagagcta gaaatagcaa gttaaaataa ggctagtccg 601 ttatcaactt gaaaagtggc accgagtcgg tgcttttttg agctccagct tttgttccct 661 ttagtgaggg ttaatttcga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa 721 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg 781 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca 841 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg 901 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg 961 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg 1021 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa 1081 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg 1141 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc 1201 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc 1261 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc 1321 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg 1381 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc 1441 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga 1501 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc 1561 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac 1621 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg 1681 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc 1741 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa 1801 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta 1861 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt 1921 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag 1981 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca 2041 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc 2101 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt 2161 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag 2221 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt 2281 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat 2341 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt 2401 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc 2461 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat 2521 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag 2581 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt 2641 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg 2701 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta 2761 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc 2821 gcgcacattt ccccgaaaag tgccacctaa attgtaagcg ttaatatttt gttaaaattc 2881 gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc 2941 ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag 3001 agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc 3061 gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa 3121 gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg 3181 aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt 3241 gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc 3301 gcgtcccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct 3361 tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg 3421 ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgagcgcgc gtaatacgac 3481 tcactatagg gcgaattggg taccgggccc cccctcgagg tcgacggtat cgataagctt 3541 ttacatccgt tgccttttcc acggtccgtg atttcatgtg cgtgcagctt caaagactgg 3601 tcgttgcgac taataagact gcagtgacag gtcgaatggt gggcaccttg ctgatgacta 3661 tctactgcaa agtctgagac aacgaacgaa acttcccaca cgaggcattt gaaactgacg 3721 gtgtctaggt aatatgcact gcaagacacg gtactggggc ctcgctgaat taggggccga 3781 tctcgttgcc ctatcagtgc tcacagtgcc gcaacgtaac accagggcag gttcttgaca 3841 gtggcaacaa tgtgcgacgg gcgtgtgaac gtttcgtagt catagcgcta gcacgtacct 3901 agccacatgg tcgtgaggag ctttaccatg cgtctagaag gtggatgcgg gacacgcctt 3961 cctggccttt ggctcccgag acgcgtgttc taaccacaaa ccttgagacg cgtgttccaa 4021 ccacgcaccc tgacacgcgt gttccaacca cgcaccctga gacgcgtgtt ctaaccacgc 4081 accctgagac gcgtgttcta accacgcacc ctgagacgcg tgttctgccg cacaatgtgc 4141 acctgtagga agctgtagtc actgctgatt ctcactgttc tcggcaaggg ccgacgaccg 4201 gagtacagtt tttgtgggca gagccgttgt gcagctttcc gttcttctcg gttgtgtcac 4261 atgtgtcatt gtcgtgtaaa cacacggttg tatgtcggtt tcgctgcacc acttcattat 4321 ttcttctggt tttttgacga gtatgcatct agacaaaatg gacaagaagt acagcatcgg 4381 cctggacatc ggcaccaact ctgtgggctg ggccgtgatc accgacgagt acaaggtgcc 4441 cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac agcatcaaga agaacctgat 4501 cggcgccctg ctgttcgaca gcggagaaac agccgaggcc acccggctga agagaaccgc 4561 cagaagaaga tacaccagac ggaagaaccg gatctgctat ctgcaagaga tcttcagcaa 4621 cgagatggcc aaggtggacg acagcttctt ccacagactg gaagagtcct tcctggtgga 4681 agaggataag aagcacgagc ggcaccccat cttcggcaac atcgtggacg aggtggccta 4741 ccacgagaag taccccacca tctaccacct gagaaagaaa ctggtggaca gcaccgacaa 4801 ggccgacctg cggctgatct atctggccct ggcccacatg atcaagttcc ggggccactt 4861 cctgatcgag ggcgacctga accccgacaa cagcgacgtg gacaagctgt tcatccagct 4921 ggtgcagacc tacaaccagc tgttcgagga aaaccccatc aacgccagcg gcgtggacgc 4981 caaggccatc ctgtctgcca gactgagcaa gagcagacgg ctggaaaatc tgatcgccca 5041 gctgcccggc gagaagaaga atggcctgtt cggcaacctg attgccctga gcctgggcct 5101 gacccccaac ttcaagagca acttcgacct ggccgaggat gccaaactgc agctgagcaa 5161 ggacacctac gacgacgacc tggacaacct gctggcccag atcggcgacc agtacgccga 5221 cctgtttctg gccgccaaga acctgtccga cgccatcctg ctgagcgaca tcctgagagt 5281 gaacaccgag atcaccaagg cccccctgag cgcctctatg atcaagagat acgacgagca 5341 ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag cagctgcctg agaagtacaa 5401 agagattttc ttcgaccaga gcaagaacgg ctacgccggc tacatcgatg gcggagccag 5461 ccaggaagag ttctacaagt tcatcaagcc catcctggaa aagatggacg gcaccgagga 5521 actgctcgtg aagctgaaca gagaggacct gctgcggaag cagcggacct tcgacaacgg 5581 cagcatcccc caccagatcc acctgggaga gctgcacgcc attctgcggc ggcaggaaga 5641 tttttaccca ttcctgaagg acaaccggga aaagatcgag aagatcctga ccttccgcat 5701 cccctactac gtgggccctc tggccagggg aaacagcaga ttcgcctgga tgaccagaaa 5761 gagcgaggaa accatcaccc cctggaactt cgaggaagtg gtggacaagg gcgccagcgc 5821 ccagagcttc atcgagcgga tgaccaactt cgataagaac ctgcccaacg agaaggtgct 5881 gcccaagcac agcctgctgt acgagtactt caccgtgtac aacgagctga ccaaagtgaa 5941 atacgtgacc gagggaatga gaaagcccgc cttcctgagc ggcgagcaga aaaaagccat 6001 cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg aagcagctga aagaggacta 6061 cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc ggcgtggaag atcggttcaa 6121 cgcctccctg ggcacatacc acgatctgct gaaaattatc aaggacaagg acttcctgga 6181 caatgaggaa aacgaggaca ttctggaaga tatcgtgctg accctgacac tgtttgagga 6241 cagagagatg atcgaggaac ggctgaaaac ctatgcccac ctgttcgacg acaaagtgat 6301 gaagcagctg aagcggcgga gatacaccgg ctggggcagg ctgagccgga agctgatcaa 6361 cggcatccgg gacaagcagt ccggcaagac aatcctggat ttcctgaagt ccgacggctt 6421 cgccaacaga aacttcatgc agctgatcca cgacgacagc ctgaccttta aagaggacat 6481 ccagaaagcc caggtgtccg gccagggcga tagcctgcac gagcacattg ccaatctggc 6541 cggcagcccc gccattaaga agggcatcct gcagacagtg aaggtggtgg acgagctcgt 6601 gaaagtgatg ggccggcaca agcccgagaa catcgtgatc gaaatggcca gagagaacca 6661 gaccacccag aagggacaga agaacagccg cgagagaatg aagcggatcg aagagggcat 6721 caaagagctg ggcagccaga tcctgaaaga acaccccgtg gaaaacaccc agctgcagaa 6781 cgagaagctg tacctgtact acctgcagaa tgggcgggat atgtacgtgg accaggaact 6841 ggacatcaac cggctgtccg actacgatgt ggaccatatc gtgcctcaga gctttctgaa 6901 ggacgactcc atcgataaca aagtgctgac tcggagcgac aagaaccggg gcaagagcga 6961 caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac tactggcgcc agctgctgaa 7021 tgccaagctg attacccaga ggaagttcga caatctgacc aaggccgaga gaggcggcct 7081 gagcgaactg gataaggccg gcttcatcaa gagacagctg gtggaaaccc ggcagatcac 7141 aaagcacgtg gcacagatcc tggactcccg gatgaacact aagtacgacg agaacgacaa 7201 actgatccgg gaagtgaaag tgatcaccct gaagtccaag ctggtgtccg atttccggaa 7261 ggatttccag ttttacaaag tgcgcgagat caacaactac caccacgccc acgacgccta 7321 cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac cctaagctgg aaagcgagtt 7381 cgtgtacggc gactacaagg tgtacgacgt gcggaagatg atcgccaaga gcgagcagga 7441 aatcggcaag gctaccgcca agtacttctt ctacagcaac atcatgaact ttttcaagac 7501 cgagattacc ctggccaacg gcgagatccg gaagcggcct ctgatcgaga caaacggcga 7561 aacaggcgag atcgtgtggg ataagggccg ggactttgcc accgtgcgga aagtgctgtc 7621 tatgccccaa gtgaatatcg tgaaaaagac cgaggtgcag acaggcggct tcagcaaaga 7681 gtctatcctg cccaagagga acagcgacaa gctgatcgcc agaaagaagg actgggaccc 7741 taagaagtac ggcggcttcg acagccccac cgtggcctat tctgtgctgg tggtggccaa 7801 agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa gagctgctgg ggatcaccat 7861 catggaaaga agcagcttcg agaagaatcc catcgacttt ctggaagcca agggctacaa 7921 agaagtgaaa aaggacctga tcatcaagct gcctaagtac tccctgttcg agctggaaaa 7981 cggccggaag agaatgctgg cctctgccgg cgaactgcag aagggaaacg aactggccct 8041 gccctccaaa tatgtgaact tcctgtacct ggccagccac tatgagaagc tgaagggctc 8101 ccccgaggat aatgagcaga aacagctgtt tgtggaacag cacaaacact acctggacga 8161 gatcatcgag cagatcagcg agttctccaa gagagtgatc ctggccgacg ctaatctgga 8221 caaggtgctg agcgcctaca acaagcacag agacaagcct atcagagagc aggccgagaa 8281 tatcatccac ctgtttaccc tgaccaatct gggagcccct gccgccttca agtactttga 8341 caccaccatc gaccggaaga ggtacaccag caccaaagag gtgctggacg ccaccctgat 8401 ccaccagagc atcaccggcc tgtacgagac acggatcgac ctgtctcagc tgggaggcga 8461 cgcctatccc tatgacgtgc ccgattatgc cagcctgggc agcggctccc ccaagaaaaa 8521 acgcaaggtg gaagatccta agaaaaagcg gaaagtggac ggcattggta gtgggagcaa 8581 cggcagcagc ggatccgtga gcaagggcga ggagctgttc accggggtgg tgcccatcct 8641 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgcgcggcg agggcgaggg 8701 cgatgccacc aacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt 8761 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc 8821 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga 8881 gcgcaccatc tccttcaagg acgacggcac ctacaagacc cgcgccgagg tgaagttcga 8941 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa 9001 catcctgggg cacaagctgg agtacaactt caacagccac aacgtctata tcacggccga 9061 caagcagaag aacggcatca aggcgaactt caagatccgc cacaacgtcg aggacggcag 9121 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct 9181 gcccgacaac cactacctga gcacccagtc caagctgagc aaagacccca acgagaagcg 9241 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga 9301 gctgtacaag tagttaatta atcaccgttg tgctcacttc tcaaatcgac aaaggaaaca 9361 cacttcgtgc agcatgtgcc ccattataaa gaaactgagt tgttccgttg tggcttgcag 9421 gtgtcacatc cacaaaaacc ggccgactct aaataggagt gtttcgcagc aagcagcgaa 9481 agtttatgac tgggtccgaa tctctgaacg gatgtgtggc ggacctggct gatgttgatc 9541 gccgtcgaca cacgcgccac atgggtcaat acacaagaca gctatcagtt gttttagtcg 9601 aaccggttaa cacaattctt gcccccccga gggggatcca ctagttctag agcggccgcc 9661 accgcggtgg agct //