质粒名称: | pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2 |
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出品公司: | 爱迪基因 |
目录编号: | 74009 |
存储实验室: | Filippo Del Bene |
载体骨架基本信息 |
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载体名称: | pminiTol2 |
载体出品公司: | -- |
载体抗性: | 氨苄青霉素 |
空载体大小: | 3459 bp |
完整质粒大小: | 9668 bp |
载体修饰: | -- |
载体类型: | CRISPR |
筛选标记: | -- |
高拷贝/低拷贝: | -- |
生长条件: | 37 ℃ |
插入基因信息 |
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插入基因名称: | 5x UAS |
物种来源: | S. cerevisiae (budding yeast) |
插入大小: | 382 bp |
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 |
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材料和方法部分: | pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2 was a gift from Filippo Del Bene (Addgene plasmid # 74009 ; http://n2t.net/addgene:74009 ; RRID:Addgene_74009) |
参考文献部分: | 2C-Cas9: a versatile tool for clonal analysis of gene function. Di Donato V, De Santis F, Auer TO, Testa N, Sanchez-Iranzo H, Mercader N, Concordet JP, Del Bene F. Genome Res. 2016 Mar 8. 10.1101/gr.196170.115 PubMed 26957310 |
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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 pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2 9671 bp ds-DNA circular SYN 10-APR-2019 DEFINITION Tissue-specific knock-out in zebrafish, Cas9 and GFP driven by a UAS promoter. ACCESSION . VERSION . KEYWORDS pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 9671) AUTHORS Di Donato V, De Santis F, Auer TO, Testa N, Sanchez-Iranzo H, Mercader N, Concordet JP, Del Bene F TITLE 2C-Cas9: a versatile tool for clonal analysis of gene function. JOURNAL Genome Res. 2016 Mar 8. PUBMED 26957310 REFERENCE 2 (bases 1 to 9671) AUTHORS . TITLE Direct Submission JOURNAL Exported Apr 10, 2019 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..9671 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 179..198 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(417..521) /gene="bla" /label=AmpR promoter rep_origin complement(599..1054) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(736..757) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 948..967 /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 1180..1202 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 1194..1211 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 1195..1211 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 1218..1237 /label=T7 /note="T7 promoter, forward primer" promoter 1218..1236 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 1895..1989 /label=5X UAS /bound_moiety="GAL4" /note="five tandem copies of the ""ScaI site"" 17-mer CGGAGTACTGTCCTCCG, an upstream activating sequence (UAS) that efficiently binds yeast Gal4 (Webster et al., 1988; Pfeiffer et al., 2010)" promoter 2008..2246 /label=hsp70 promoter /note="Drosophila melanogaster hsp70Bb promoter" primer_bind 2193..2214 /label=pCasper-hs /note="Drosophila Hsp70 promoter, forward primer" regulatory 2261..2270 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2279..2299 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 2306..6406 /codon_start=1 /product="Cas9 (Csn1) endonuclease from the Streptococcus pyogenes MGAS15252 Type II CRISPR/Cas system" /label=Cas9 /note="generates RNA-guided double strand breaks in DNA" /translation="MDKKYSIGLDIGTNSVGWAVITDDYKVPSKKFKVLGNTDRHSIKK NLIGALLFGSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLADSTDKADLRLIYLALAHMIK FRGHFLIEGDLNPDNSDVDKLFIQLVQIYNQLFEENPINASRVDAKAILSARLSKSRRL ENLIAQLPGEKRNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ IGDQYADLFLAAKNLSDAILLSDILRVNSEITKAPLSASMIKRYDEHHQDLTLLKALVR QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD SVEISGVEDRFNASLGAYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDRGMIEER LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM QLIHDDSLTFKEDIQKAQVSGQGHSLHEQIANLAGSPAIKKGILQTVKIVDELVKVMGH KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFIKDDSIDNKVLTRSDKNRGKSDNVPS EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD ATLIHQSITGLYETRIDLSQLGGD" RBS 6319..6327 /label=Shine-Dalgarno sequence /note="full consensus sequence for ribosome-binding sites upstream of start codons in E. coli; complementary to a region in the 3' end of the 16S rRNA (Chen et al., 1994)" CDS 6422..6442 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 6458..6511 /codon_start=1 /product="2A peptide from Thosea asigna virus capsid protein" /label=T2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="EGRGSLLTCGDVEENPGP" CDS 6512..7231 /codon_start=1 /product="enhanced GFP" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(6557..6578) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(6818..6837) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 7165..7186 /label=EGFP-C /note="EGFP, forward primer" promoter complement(7239..7256) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(7285..7304) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 7339..7358 /label=SV40pA-R /note="SV40 polyA, reverse primer" promoter complement(8030..8048) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(8031..8048) /label=SP6 /note="SP6 promoter, forward primer" primer_bind complement(8066..8082) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(8066..8082) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(8079..8101) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 8090..8106 /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(8114..8144) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 8159..8180 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(8297..8314) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(8468..9056) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(8548..8567) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(9227..416) /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" ORIGIN 1 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat 61 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag 121 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat 181 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg 241 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca 301 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga 361 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc 421 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata 481 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg 541 ccacctgatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggaa 601 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt 661 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata 721 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac 781 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa 841 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc 901 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg 961 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca 1021 cccgccgcgc ttaatgcgcc gctacagggc gcgtccattc gccattcagg ctgcgcaact 1081 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat 1141 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa 1201 cgacggccag tgaattgtaa tacgactcac tatagggcga attgggccca gaggtgtaaa 1261 gtacttgagt aattttactt gattactgta cttaagtatt atttttgggg atttttactt 1321 tacttgagta caattaaaaa tcaatacttt tacttttact taattacatt tttttagaaa 1381 aaaaagtact ttttactcct tacaatttta tttacagtca aaaagtactt attttttgga 1441 gatcacttca ttctattttc ccttgctatt accaaaccaa ttgaattgcg ctgatgccca 1501 gtttaattta aatagatccc atgggtcgac tacgcttaca gctccaaaac tctacggtaa 1561 acctacataa actgctggtt ttcaaatttt aaagaattta agggtttaca ggtttactac 1621 tacacagtga tttactgaca catgtaagtg taaatgagtt gaataagtaa gtaagccata 1681 taccacacat gaaacacata cccagaagtc actggtatat atagccgtcc tccagactcc 1741 cagctcgaga gaccgagaga gggtctcagt ttaagagcta tgctggaaac agcatagcaa 1801 gtttaaataa ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgctttttt 1861 tggtcgacgg atccaagctt gcatgcctgc aggtcggagt actgtcctcc gagcggagta 1921 ctgtcctccg agcggagtac tgtcctccga gcggagtact gtcctccgag cggagtactg 1981 tcctccgagc ggagactcta gcgagcgccg gagtataaat agaggcgctt cgtctacgga 2041 gcgacaattc aattcaaaca agcaaagtga acacgtcgct aagcgaaagc taagcaaata 2101 aacaagcgca gctgaacaag ctaaacaatc tgcagtaaag tgcaagttaa agtgaatcaa 2161 ttaaaagtaa ccagcaacca agtaaatcaa ctgcaactac tgaaatctgc caagaagtaa 2221 ttattgaata caagaagaga actctgaata gggaattggg gccaccatgg cttctccacc 2281 taagaagaag agaaaggtgg gaagcatgga taagaagtat agcatcggcc tggatattgg 2341 aactaactcc gtgggttggg cagtgattac agacgactac aaggtcccta gcaagaaatt 2401 taaggtgctg ggtaacaccg acaggcacag catcaagaaa aatctgattg gagccctgct 2461 gttcggttct ggagagactg ccgaagcaac acgcctgaaa agaacagcaa gaaggcgcta 2521 taccagaagg aagaatagaa tctgttacct gcaggagatt ttctctaacg aaatggctaa 2581 ggtggacgat tcattctttc ataggctgga ggaaagtttc ctggtcgagg aagataagaa 2641 acacgagcgc catcctatct ttggaaacat tgtggacgag gtcgcctatc acgaaaaata 2701 cccaaccatc tatcatctgc gcaagaaact ggctgactct actgataaag ccgacctgag 2761 actgatctat ctggctctgg cccacatgat taagttcagg ggtcattttc tgatcgaggg 2821 cgatctgaac cccgacaatt ccgatgtgga caagctgttc atccagctgg tccagattta 2881 caatcagctg tttgaggaaa accctattaa tgcttccaga gtggacgcaa aagctatcct 2941 gtcagccagg ctgtccaagt cacgcagact ggagaacctg attgcacagc tgcccggaga 3001 aaagaggaac ggtctgtttg gaaatctgat cgctctgagt ctgggcctga ctcctaactt 3061 caaaagcaat tttgatctgg ctgaggacgc caaactgcag ctgtcaaagg acacatatga 3121 cgatgacctg gataacctgc tggcacagat cggagatcag tacgctgacc tgttcctggc 3181 tgccaaaaat ctgtccgacg caatcctgct gtcagatatt ctgagagtga acagcgagat 3241 tacaaaagct cctctgagtg ccagcatgat caagagatat gacgagcacc atcaggatct 3301 gaccctgctg aaggctctgg tcaggcagca gctgccagag aagtacaagg aaattttctt 3361 tgatcagtcc aagaacggct acgccggtta tatcgacgga ggcgcatcac aggaggaatt 3421 ctacaagttt atcaaaccta ttctggagaa gatggacgga actgaggaac tgctggtgaa 3481 actgaataga gaggacctgc tgaggaagca gcgcacattt gataacggtt ccatcccaca 3541 ccagattcat ctgggagagc tgcacgctat cctgaggcgc caggaagact tctacccctt 3601 tctgaaagat aaccgcgaga agatcgaaaa aattctgacc ttcagaatcc cttactatgt 3661 gggtccactg gctcgcggaa acagcagatt tgcctggatg actcgcaaat ccgaggaaac 3721 cattactcct tggaacttcg aggaagtggt cgataagggc gcctctgcac agtccttcat 3781 cgagagaatg actaattttg acaaaaacct gcccaatgag aaagtgctgc ctaagcactc 3841 cctgctgtac gagtatttca ctgtctataa cgaactgaca aaggtgaaat acgtcaccga 3901 gggcatgaga aagccagcct tcctgtcagg agagcagaag aaagcaatcg tggatctgct 3961 gtttaaaacc aacaggaaag tgactgtcaa gcagctgaag gaggactact tcaagaaaat 4021 tgaatgcttc gattccgtgg agatcagcgg agtcgaagac agatttaacg caagcctggg 4081 cgcttaccac gatctgctga agatcattaa ggataaagac ttcctggaca acgaggaaaa 4141 tgaggatatc ctggaagaca ttgtgctgac actgaccctg tttgaggaca gaggaatgat 4201 cgaggaaaga ctgaaaacct atgctcatct gttcgatgac aaggtgatga aacagctgaa 4261 gagaaggcgc tacactggct ggggtagact gagcaggaag ctgatcaacg gcattaggga 4321 taaacagtca ggaaagacaa tcctggactt tctgaaaagt gatggcttcg ccaaccgcaa 4381 ttttatgcag ctgattcacg atgacagtct gaccttcaaa gaggacatcc agaaggctca 4441 ggtgtctgga cagggccact ccctgcatga gcagattgca aacctggctg gaagcccagc 4501 catcaagaaa ggcattctgc agacagtgaa aatcgtcgat gagctggtga aagtcatggg 4561 ccataagccc gaaaacatcg tgattgagat ggctcgcgaa aatcagacaa cccagaaggg 4621 tcagaagaac agtagagaga ggatgaaaag aatcgaggaa ggcattaagg agctgggtag 4681 ccagatcctg aaagagcacc cagtggaaaa cacacagctg cagaatgaga agctgtatct 4741 gtactatctg cagaatggaa gagatatgta cgtggaccag gagctggata ttaacaggct 4801 gtctgattac gacgtggatc atatcgtccc ccagagtttc atcaaagatg acagcattga 4861 caacaaggtg ctgaccaggt ccgacaaaaa cagaggaaaa tcagataatg tccctagtga 4921 ggaagtggtc aagaaaatga agaactactg gagacagctg ctgaatgcca aactgatcac 4981 tcagaggaag tttgataacc tgacaaaagc agagcgcggt ggactgtcag aactggacaa 5041 agctggattc atcaagaggc agctggtgga aacacgccag atcactaaac acgtcgcaca 5101 gattctggat agtcgcatga acacaaagta cgatgagaat gacaaactga tcagagaagt 5161 gaaggtcatt accctgaaga gtaaactggt cagcgacttt aggaaagatt tccagtttta 5221 taaggtccgc gagattaaca attatcacca tgcccatgac gcatacctga acgccgtggt 5281 cggtaccgca ctgatcaaga aatacccaaa actggagagc gaattcgtgt acggagacta 5341 taaggtgtac gatgtcagaa aaatgatcgc caagtccgag caggaaattg gaaaagctac 5401 tgccaagtat ttcttttact caaacatcat gaatttcttt aagacagaga tcaccctggc 5461 caatggagaa atccgcaaaa ggcccctgat tgagacaaac ggagagacag gcgaaatcgt 5521 gtgggacaaa ggcagagatt ttgcaaccgt gaggaaggtc ctgagcatgc ctcaagtgaa 5581 tatcgtcaag aaaactgagg tgcagacagg cggtttctca aaagaaagta ttctgccaaa 5641 acgcaactct gataagctga tcgctagaaa gaaagactgg gaccctaaga agtatggagg 5701 ctttgactct cccactgtgg catactccgt cctggtggtc gctaaggtgg agaagggcaa 5761 aagcaagaaa ctgaaatctg tcaaggagct gctgggtatc acaattatgg agagaagctc 5821 tttcgagaag aacccaatcg attttctgga ggccaaaggt tataaggaag tgaagaaaga 5881 cctgatcatt aaactgccca agtacagtct gtttgagctg gaaaacggca ggaaacgcat 5941 gctggcaagc gctggagagc tgcagaaagg caatgaactg gccctgcctt ctaagtacgt 6001 gaacttcctg tatctggcaa gccactacga gaagctgaaa ggatctccag aggataacga 6061 acagaaacag ctgtttgtgg agcagcacaa gcattatctg gacgagatca ttgaacagat 6121 tagcgagttc tctaaaagag tgatcctggc cgacgcaaat ctggataagg tcctgtctgc 6181 ttacaacaaa cacagagata agcccatcag ggagcaggcc gaaaatatca ttcatctgtt 6241 cactctgaca aacctgggcg cacctgcagc tttcaagtac ttcgacacta caatcgatag 6301 aaagaggtac acctccacta aggaggtgct ggacgctaca ctgatccatc agagtattac 6361 cggcctgtac gaaacaagga ttgacctgtc tcagctgggt ggcgactcac cggtgagatc 6421 tcctaagaag aagagaaagg tgaggtccgg cggcggagag ggcagaggaa gtcttctaac 6481 atgcggtgac gtggaggaga atcccggccc aatggtgagc aagggcgagg agctgttcac 6541 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt 6601 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac 6661 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca 6721 gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc 6781 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg 6841 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga 6901 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa 6961 cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca 7021 caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg 7081 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa 7141 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat 7201 cactctcggc atggacgagc tgtacaagta atctagaact atagtgagtc gtattacgta 7261 gatccagaca tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga 7321 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc 7381 tgcaataaac cttgtacaaa gtgggggatc ctacgcttac agctccaaaa ctctacggta 7441 aacctacata aactgctggt tttcaaattt taaagaattt aagggtttac aggtttacta 7501 ctacacagtg atttactgac acatgtaagt gtaaatgagt tgaataagta agtaagccat 7561 ataccacaca tgaaacacat acccagaagt cactggtata tatagccgtc ctccagactc 7621 ccagctcgag acgattagat ctaatgcgtc tcgtttaaga gctatgctgg aaacagcata 7681 gcaagtttaa ataaggctag tccgttatca acttgaaaaa gtggcaccga gtcggtgctt 7741 tttttcaatg atgaattaat tcgagctcca ccgcggaagc ttaaacaaga atctctagtt 7801 ttctttcttg cttttacttt tacttcctta atactcaagt acaattttaa tggagtactt 7861 ttttactttt actcaagtaa gattctagcc agatactttt acttttaatt gagtaaaatt 7921 ttccctaagt acttgtactt tcacttgagt aaaatttttg agtacttttt acacctctgc 7981 tcgaccatat gggagagctc ccaacgcgtt ggatgcatag cttgagtatt ctatagtgtc 8041 acctaaatag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc 8101 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat 8161 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc 8221 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg 8281 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag 8341 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag 8401 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc 8461 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc 8521 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc 8581 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt 8641 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg 8701 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat 8761 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag 8821 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt 8881 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc 8941 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta 9001 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag 9061 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggca 9121 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa 9181 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa 9241 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc 9301 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga 9361 taccgcgaga tccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa 9421 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt 9481 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg 9541 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc 9601 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg 9661 gtcctccgat c //
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