质粒名称: | pDIRECT_23A |
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出品公司: | 爱迪基因 |
目录编号: | 91138 |
存储实验室: | Daniel Voytas |
载体骨架基本信息 |
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载体名称: | pTRANS_230 |
载体出品公司: | -- |
载体抗性: | 卡那霉素 |
空载体大小: | -- |
完整质粒大小: | -- |
载体修饰: | -- |
载体类型: | 植物表达,CRISPR |
筛选标记: | Basta |
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: | Engineering Reagent: 35S:AtCas9 + AtU6:gRNA |
物种来源: | Synthetic |
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 |
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材料和方法部分: | pDIRECT_23A was a gift from Daniel Voytas (Addgene plasmid # 91138 ; http://n2t.net/addgene:91138 ; RRID:Addgene_91138) |
参考文献部分: | A multi-purpose toolkit to enable advanced genome engineering in plants. Cermak T, Curtin SJ, Gil-Humanes J, Cegan R, Kono TJY, Konecna E, Belanto JJ, Starker CG, Mathre JW, Greenstein RL, Voytas DF. Plant Cell. 2017 May 18. pii: tpc.00922.2016. doi: 10.1105/tpc.16.00922. 10.1105/tpc.16.00922 PubMed 28522548 |
买家导航
pDIRECT23A载体基本信息
pDIRECT23A载体图谱质粒图谱和多克隆位点信息
pDIRECT23A载体简介
pDIRECT23A载体序列
LOCUS pDIRECT_23A 14676 bp ds-DNA circular SYN 06-AUG-2018 DEFINITION Direct Cloning, Type: T-DNA, Engineering Reagent: 35S:AtCas9 + AtU6:gRNA, Plant Selection: 2x35S:bar. ACCESSION . VERSION . KEYWORDS pDIRECT_23A SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 14676) AUTHORS Cermak T, Curtin SJ, Gil-Humanes J, Cegan R, Kono TJY, Konecna E, Belanto JJ, Starker CG, Mathre JW, Greenstein RL, Voytas DF TITLE A multi-purpose toolkit to enable advanced genome engineering in plants. JOURNAL Plant Cell. 2017 May 18. pii: tpc.00922.2016. doi: 10.1105/tpc.16.00922. PUBMED 28522548 REFERENCE 2 (bases 1 to 14676) AUTHORS . TITLE Direct Submission JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..14676 /organism="synthetic DNA construct" /mol_type="other DNA" CDS complement(291..1085) /codon_start=1 /gene="aphA-3" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin" /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF" misc_feature 1510..1534 /label=LB T-DNA repeat /note="left border repeat from nopaline C58 T-DNA" polyA_signal 1612..1786 /label=CaMV poly(A) signal /note="cauliflower mosaic virus polyadenylation signal" CDS complement(1793..2344) /codon_start=1 /gene="Streptomyces hygroscopicus bar" /product="phosphinothricin acetyltransferase" /label=BlpR /note="confers resistance to bialophos or phosphinothricin" /translation="MSPERRPADIRRATEADMPAVCTIVNHYIETSTVNFRTEPQEPQE WTDDLVRLRERYPWLVAEVDGEVAGIAYAGPWKARNAYDWTAESTVYVSPRHQRTGLGS TLYTHLLKSLEAQGFKSVVAVIGLPNDPSVRMHEALGYAPRGMLRAAGFKHGNWHDVGF WQLDFSLPVPPRPVLPVTEI" promoter complement(2389..3065) /label=CaMV 35S promoter (enhanced) /note="cauliflower mosaic virus 35S promoter with a duplicated enhancer region" protein_bind 3256..3277 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 3292..3322 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 3330..3346 /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 3354..3370 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" promoter 3900..4245 /label=CaMV 35S promoter /note="strong constitutive promoter from cauliflower mosaic virus" CDS 4279..8382 /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 8395..8415 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS complement(9141..9446) /codon_start=1 /gene="lacZ fragment" /product="LacZ-alpha fragment of beta-galactosidase" /label=lacZ-alpha /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGTCL" primer_bind 9311..9327 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 9337..9356 /label=T7 /note="T7 promoter, forward primer" promoter 9337..9355 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(9403..9423) /label=T3 /note="T3 promoter, forward primer" promoter complement(9403..9421) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(9442..9458) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" protein_bind 9466..9482 /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(9490..9520) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 9535..9556 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." misc_RNA 9595..9670 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" primer_bind complement(9687..9703) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" misc_feature 9906..9930 /label=RB T-DNA repeat /note="right border repeat from nopaline C58 T-DNA" CDS 11231..11860 /codon_start=1 /product="stability protein from the Pseudomonas plasmid pVS1 (Heeb et al., 2000)" /label=pVS1 StaA /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI" CDS 12289..13362 /codon_start=1 /product="replication protein from the Pseudomonas plasmid pVS1 (Heeb et al., 2000)" /label=pVS1 RepA /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI LVMRYRNLIEGEASAGS" rep_origin 13428..13622 /label=pVS1 oriV /note="origin of replication for the Pseudomonas plasmid pVS1 (Heeb et al., 2000)" primer_bind complement(13858..13880) /label=pGEX 3' /note="pGEX vectors, reverse primer" misc_feature 13966..14106 /label=bom /note="basis of mobility region from pBR322" primer_bind 14021..14040 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind complement(14121..14138) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(14292..204) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(14372..14391) /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca 61 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct 121 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt 181 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct 241 cagtggaacg aaaactcacg ttaagggatt ttggtcatgc attctaggta ctaaaacaat 301 tcatccagta aaatataata ttttattttc tcccaatcag gcttgatccc cagtaagtca 361 aaaaatagct cgacatactg ttcttccccg atatcctccc tgatcgaccg gacgcagaag 421 gcaatgtcat accacttgtc cgccctgccg cttctcccaa gatcaataaa gccacttact 481 ttgccatctt tcacaaagat gttgctgtct cccaggtcgc cgtgggaaaa gacaagttcc 541 tcttcgggct tttccgtctt taaaaaatca tacagctcgc gcggatcttt aaatggagtg 601 tcttcttccc agttttcgca atccacatcg gccagatcgt tattcagtaa gtaatccaat 661 tcggctaagc ggctgtctaa gctattcgta tagggacaat ccgatatgtc gatggagtga 721 aagagcctga tgcactccgc atacagctcg ataatctttt cagggctttg ttcatcttca 781 tactcttccg agcaaaggac gccatcggcc tcactcatga gcagattgct ccagccatca 841 tgccgttcaa agtgcaggac ctttggaaca ggcagctttc cttccagcca tagcatcatg 901 tccttttccc gttccacatc ataggtggtc cctttatacc ggctgtccgt catttttaaa 961 tataggtttt cattttctcc caccagctta tataccttag caggagacat tccttccgta 1021 tcttttacgc agcggtattt ttcgatcagt tttttcaatt ccggtgatat tctcatttta 1081 gccatttatt atttccttcc tcttttctac agtatttaaa gataccccaa gaagctaatt 1141 ataacaagac gaactccaat tcactgttcc ttgcattcta aaaccttaaa taccagaaaa 1201 cagctttttc aaagttgttt tcaaagttgg cgtataacat agtatcgacg gagccgattt 1261 tgaaaccgcg gtgatcacag gcagcaacgc tctgtcatcg ttacaatcaa catgctaccc 1321 tccgcgagat catccgtgtt tcaaacccgg cagcttagtt gccgttcttc cgaatagcat 1381 cggtaacatg agcaaagtct gccgccttac aacggctctc ccgctgacgc cgtcccggac 1441 tgatgggctg cctgtatcga gtggtgattt tgtgccgagc tgccggtcgg ggagctgttg 1501 gctggctggt ggcaggatat attgtggtgt aaacaaattg acgcttagac aacttaataa 1561 cacattgcgg acgtttttaa tgtactgaat taacgccgaa ttaattcggg ggatctggat 1621 tttagtactg gattttggtt ttaggaatta gaaattttat tgatagaagt attttacaaa 1681 tacaaataca tactaagggt ttcttatatg ctcaacacat gagcgaaacc ctataggaac 1741 cctaattccc ttatctggga actactcaca cattattatg gagaaactcg agtcaaatct 1801 cggtgacggg caggaccgga cggggcggta ccggcaggct gaagtccagc tgccagaaac 1861 ccacgtcatg ccagttcccg tgcttgaagc cggccgcccg cagcatgccg cggggggcat 1921 atccgagcgc ctcgtgcatg cgcacgctcg ggtcgttggg cagcccgatg acagcgacca 1981 cgctcttgaa gccctgtgcc tccagggact tcagcagatg ggtgtagagc gtggagccca 2041 gtcccgtccg ctggtggcgg ggggagacgt acacggtcga ctcggccgtc cagtcgtagg 2101 cgttgcgtgc cttccagggg cccgcgtagg cgatgccggc gacctcgccg tccacctcgg 2161 cgacgagcca gggatagcgc tcccgcagac ggacgaggtc gtccgtccac tcctgcggtt 2221 cctgcggctc ggtacggaag ttgaccgtgc ttgtctcgat gtagtggttg acgatggtgc 2281 agaccgccgg catgtccgcc tcggtggcac ggcggatgtc ggccgggcgt cgttctgggc 2341 tcatggtaga ctcgagagag atagatttgt agagagagac tggtgatttc agcgtgtcct 2401 ctccaaatga aatgaacttc cttatataga ggaagggtct tgcgaaggat agtgggattg 2461 tgcgtcatcc cttacgtcag tggagatatc acatcaatcc acttgctttg aagacgtggt 2521 tggaacgtct tctttttcca cgatgctcct cgtgggtggg ggtccatctt tgggaccact 2581 gtcggcagag gcatcttgaa cgatagcctt tcctttatcg caatgatggc atttgtaggt 2641 gccaccttcc ttttctactg tccttttgat gaagtgacag atagctgggc aatggaatcc 2701 gaggaggttt cccgatatta ccctttgttg aaaagtctca atagcccttt ggtcttctga 2761 gactgtatct ttgatattct tggagtagac gagagtgtcg tgctccacca tgttcacatc 2821 aatccacttg ctttgaagac gtggttggaa cgtcttcttt ttccacgatg ctcctcgtgg 2881 gtgggggtcc atctttggga ccactgtcgg cagaggcatc ttgaacgata gcctttcctt 2941 tatcgcaatg atggcatttg taggtgccac cttccttttc tactgtcctt ttgatgaagt 3001 gacagatagc tgggcaatgg aatccgagga ggtttcccga tattaccctt tgttgaaaag 3061 tctcaatagc cctttggtct tctgagactg tatctttgat attcttggag tagacgagag 3121 tgtcgtgctc caccatgttg gcaagctgct ctagccaata cgcaaaccgc ctctccccgc 3181 gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag 3241 tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt 3301 tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa 3361 cagctatgac atgattacga attcgagctc ggtacccggg gatcggcgcg ccagatttgc 3421 cttttcaatt tcagaaagaa tgctaaccca cagatggtta gagaggctta cgcagcaggt 3481 atcatcaaga cgatctaccc gagcaataat ctccaggaaa tcaaatacct tcccaagaag 3541 gttaaagatg cagtcaaaag attcaggact aactgcatca agaacacaga gaaagatata 3601 tttctcaaga tcagaagtac tattccagta tggacgattc aaggcttgct tcacaaacca 3661 aggcaagtaa tagagattgg agtctctaaa aaggtagttc ccactgaatc aaaggccatg 3721 gagtcaaaga ttcaaataga ggacctaaca gaactcgccg taaagactgg cgaacagttc 3781 atacagagtc tcttacgact caatgacaag aagaaaatct tcgtcaacat ggtggagcac 3841 gacacacttg tctactccaa aaatatcaaa gatacagtct cagaagacca aagggcaatt 3901 gagacttttc aacaaagggt aatatccgga aacctcctcg gattccattg cccagctatc 3961 tgtcacttta ttgtgaagat agtggaaaag gaaggtggct cctacaaatg ccatcattgc 4021 gataaaggaa aggccatcgt tgaagatgcc tctgccgaca gtggtcccaa agatggaccc 4081 ccacccacga ggagcatcgt ggaaaaagaa gacgttccaa ccacgtcttc aaagcaagtg 4141 gattgatgtg atatctccac tgacgtaagg gatgacgcac aatcccacta tccttcgcaa 4201 gacccttcct ctatataagg aagttcattt catttggaga gaacacgggg gactcctgca 4261 ggtagatcgc tcgtcgacat ggataagaag tactctatcg gactcgatat cggaactaac 4321 tctgtgggat gggctgtgat caccgatgag tacaaggtgc catctaagaa gttcaaggtt 4381 ctcggaaaca ccgataggca ctctatcaag aaaaacctta tcggtgctct cctcttcgat 4441 tctggtgaaa ctgctgaggc taccagactc aagagaaccg ctagaagaag gtacaccaga 4501 agaaagaaca ggatctgcta cctccaagag atcttctcta acgagatggc taaagtggat 4561 gattcattct tccacaggct cgaagagtca ttcctcgtgg aagaagataa gaagcacgag 4621 aggcacccta tcttcggaaa catcgttgat gaggtggcat accacgagaa gtaccctact 4681 atctaccacc tcagaaagaa gctcgttgat tctactgata aggctgatct caggctcatc 4741 tacctcgctc tcgctcacat gatcaagttc agaggacact tcctcatcga gggtgatctc 4801 aaccctgata actctgatgt ggataagttg ttcatccagc tcgtgcagac ctacaaccag 4861 cttttcgaag agaaccctat caacgcttca ggtgtggatg ctaaggctat cctctctgct 4921 aggctctcta agtcaagaag gcttgagaac ctcattgctc agctccctgg tgagaagaag 4981 aacggacttt tcggaaactt gatcgctctc tctctcggac tcacccctaa cttcaagtct 5041 aacttcgatc tcgctgagga tgcaaagctc cagctctcaa aggataccta cgatgatgat 5101 ctcgataacc tcctcgctca gatcggagat cagtacgctg atttgttcct cgctgctaag 5161 aacctctctg atgctatcct cctcagtgat atcctcagag tgaacaccga gatcaccaag 5221 gctccactct cagcttctat gatcaagaga tacgatgagc accaccagga tctcacactt 5281 ctcaaggctc ttgttagaca gcagctccca gagaagtaca aagagatttt cttcgatcag 5341 tctaagaacg gatacgctgg ttacatcgat ggtggtgcat ctcaagaaga gttctacaag 5401 ttcatcaagc ctatcctcga gaagatggat ggaaccgagg aactcctcgt gaagctcaat 5461 agagaggatc ttctcagaaa gcagaggacc ttcgataacg gatctatccc tcatcagatc 5521 cacctcggag agttgcacgc tatccttaga aggcaagagg atttctaccc attcctcaag 5581 gataacaggg aaaagattga gaagattctc accttcagaa tcccttacta cgtgggacct 5641 ctcgctagag gaaactcaag attcgcttgg atgaccagaa agtctgagga aaccatcacc 5701 ccttggaact tcgaagaggt ggtggataag ggtgctagtg ctcagtcttt catcgagagg 5761 atgaccaact tcgataagaa ccttccaaac gagaaggtgc tccctaagca ctctttgctc 5821 tacgagtact tcaccgtgta caacgagttg accaaggtta agtacgtgac cgagggaatg 5881 aggaagcctg cttttttgtc aggtgagcaa aagaaggcta tcgttgatct cttgttcaag 5941 accaacagaa aggtgaccgt gaagcagctc aaagaggatt acttcaagaa aatcgagtgc 6001 ttcgattcag ttgagatttc tggtgttgag gataggttca acgcatctct cggaacctac 6061 cacgatctcc tcaagatcat taaggataag gatttcttgg ataacgagga aaacgaggat 6121 atcttggagg atatcgttct taccctcacc ctctttgaag atagagagat gattgaagaa 6181 aggctcaaga cctacgctca tctcttcgat gataaggtga tgaagcagtt gaagagaaga 6241 agatacactg gttggggaag gctctcaaga aagctcatta acggaatcag ggataagcag 6301 tctggaaaga caatccttga tttcctcaag tctgatggat tcgctaacag aaacttcatg 6361 cagctcatcc acgatgattc tctcaccttt aaagaggata tccagaaggc tcaggtttca 6421 ggacagggtg atagtctcca tgagcatatc gctaacctcg ctggatctcc tgcaatcaag 6481 aagggaatcc tccagactgt gaaggttgtg gatgagttgg tgaaggtgat gggaaggcat 6541 aagcctgaga acatcgtgat cgaaatggct agagagaacc agaccactca gaagggacag 6601 aagaactcta gggaaaggat gaagaggatc gaggaaggta tcaaagagct tggatctcag 6661 atcctcaaag agcaccctgt tgagaacact cagctccaga atgagaagct ctacctctac 6721 tacctccaga acggaaggga tatgtatgtg gatcaagagt tggatatcaa caggctctct 6781 gattacgatg ttgatcatat cgtgccacag tcattcttga aggatgattc tatcgataac 6841 aaggtgctca ccaggtctga taagaacagg ggtaagagtg ataacgtgcc aagtgaagag 6901 gttgtgaaga aaatgaagaa ctattggagg cagctcctca acgctaagct catcactcag 6961 agaaagttcg ataacttgac taaggctgag aggggaggac tctctgaatt ggataaggca 7021 ggattcatca agaggcagct tgtggaaacc aggcagatca ctaagcacgt tgcacagatc 7081 ctcgattcta ggatgaacac caagtacgat gagaacgata agttgatcag ggaagtgaag 7141 gttatcaccc tcaagtcaaa gctcgtgtct gatttcagaa aggatttcca attctacaag 7201 gtgagggaaa tcaacaacta ccaccacgct cacgatgctt accttaacgc tgttgttgga 7261 accgctctca tcaagaagta tcctaagctc gagtcagagt tcgtgtacgg tgattacaag 7321 gtgtacgatg tgaggaagat gatcgctaag tctgagcaag agatcggaaa ggctaccgct 7381 aagtatttct tctactctaa catcatgaat ttcttcaaga ccgagattac cctcgctaac 7441 ggtgagatca gaaagaggcc actcatcgag acaaacggtg aaacaggtga gatcgtgtgg 7501 gataagggaa gggatttcgc taccgttaga aaggtgctct ctatgccaca ggtgaacatc 7561 gttaagaaaa ccgaggtgca gaccggtgga ttctctaaag agtctatcct ccctaagagg 7621 aactctgata agctcattgc taggaagaag gattgggacc ctaagaaata cggtggtttc 7681 gattctccta ccgtggctta ctctgttctc gttgtggcta aggttgagaa gggaaagagt 7741 aagaagctca agtctgttaa ggaacttctc ggaatcacta tcatggaaag gtcatctttc 7801 gagaagaacc caatcgattt cctcgaggct aagggataca aagaggttaa gaaggatctc 7861 atcatcaagc tcccaaagta ctcactcttc gaactcgaga acggtagaaa gaggatgctc 7921 gcttctgctg gtgagcttca aaagggaaac gagcttgctc tcccatctaa gtacgttaac 7981 tttctttacc tcgcttctca ctacgagaag ttgaagggat ctccagaaga taacgagcag 8041 aagcaacttt tcgttgagca gcacaagcac tacttggatg agatcatcga gcagatctct 8101 gagttctcta aaagggtgat cctcgctgat gcaaacctcg ataaggtgtt gtctgcttac 8161 aacaagcaca gagataagcc tatcagggaa caggcagaga acatcatcca tctcttcacc 8221 cttaccaacc tcggtgctcc tgctgctttc aagtacttcg atacaaccat cgataggaag 8281 agatacacct ctaccaaaga agtgctcgat gctaccctca tccatcagtc tatcactgga 8341 ctctacgaga ctaggatcga tctctcacag ctcggtggtg attcaagggc tgatcctaag 8401 aagaagagga aggtttgact cgagatatga agatgaagat gaaatatttg gtgtgtcaaa 8461 taaaaagctt gtgtgcttaa gtttgtgttt ttttcttggc ttgttgtgtt atgaatttgt 8521 ggctttttct aatattaaat gaatgtaaga tcacattata atgaataaac aaatgtttct 8581 ataatccatt gtgaatgttt tgttggatct cttctgcagc atataactac tgtatgtgct 8641 atggtatgga ctatggaata tgattaaaga taaggagctc cggtgacgga cccatggctt 8701 cgttgaacaa cggaaactcg acttgccttc cgcacaatac atcatttctt cttagctttt 8761 tttcttcttc ttcgttcata cagttttttt ttgtttatca gcttacattt tcttgaaccg 8821 tagctttcgt tttcttcttt ttaactttcc attcggagtt tttgtatctt gtttcatagt 8881 ttgtcccagg attagaatga ttaggcatcg aaccttcaag aatttgattg aataaaacat 8941 cttcattctt aagatatgaa gataatcttc aaaaggcccc tgggaatctg aaagaagaga 9001 agcaggccca tttatatggg aaagaacaat agtatttctt atataggccc atttaagttg 9061 aaaacaatct tcaaaagtcc cacatcgctt agataagaaa acgaagctga gtttatatac 9121 agctagagtc gaagtagtga ttagaggcag gtgccgctac agggcgcgtc ccattcgcca 9181 ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag 9241 ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag 9301 tcacgacgtt gtaaaacgac ggccagtgag cgcgcgtaat acgactcact atagggcgaa 9361 ttgggtaccg ggccccccct cgaggtcctc cagcttttgt tccctttagt gagggttaat 9421 tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac 9481 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt 9541 gagctaactc acattaattg cgttgcgctc actgcccgct ttccacctgc ggtcgtttta 9601 gagctagaaa tagcaagtta aaataaggct agtccgttat caacttgaaa aagtggcacc 9661 gagtcggtgc tttttttccc ggtggcactg gccgtcgttt tacaacgtcg tgactgggaa 9721 aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt 9781 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa 9841 tgctagagca gcttgagctt ggatcagatt gtcgtttccc gccttcagtt taaactatca 9901 gtgtttgaca ggatatattg gcgggtaaac ctaagagaaa agagcgttta ttagaataat 9961 cggatattta aaagggcgtg aaaaggttta tccgttcgtc catttgtatg tgcatgccaa 10021 ccacagggtt cccctcggga tcaaagtact ttgatccaac ccctccgctg ctatagtgca 10081 gtcggcttct gacgttcagt gcagccgtct tctgaaaacg acatgtcgca caagtcctaa 10141 gttacgcgac aggctgccgc cctgcccttt tcctggcgtt ttcttgtcgc gtgttttagt 10201 cgcataaagt agaatacttg cgactagaac cggagacatt acgccatgaa caagagcgcc 10261 gccgctggcc tgctgggcta tgcccgcgtc agcaccgacg accaggactt gaccaaccaa 10321 cgggccgaac tgcacgcggc cggctgcacc aagctgtttt ccgagaagat caccggcacc 10381 aggcgcgacc gcccggagct ggccaggatg cttgaccacc tacgccctgg cgacgttgtg 10441 acagtgacca ggctagaccg cctggcccgc agcacccgcg acctactgga cattgccgag 10501 cgcatccagg aggccggcgc gggcctgcgt agcctggcag agccgtgggc cgacaccacc 10561 acgccggccg gccgcatggt gttgaccgtg ttcgccggca ttgccgagtt cgagcgttcc 10621 ctaatcatcg accgcacccg gagcgggcgc gaggccgcca aggcccgagg cgtgaagttt 10681 ggcccccgcc ctaccctcac cccggcacag atcgcgcacg cccgcgagct gatcgaccag 10741 gaaggccgca ccgtgaaaga ggcggctgca ctgcttggcg tgcatcgctc gaccctgtac 10801 cgcgcacttg agcgcagcga ggaagtgacg cccaccgagg ccaggcggcg cggtgccttc 10861 cgtgaggacg cattgaccga ggccgacgcc ctggcggccg ccgagaatga acgccaagag 10921 gaacaagcat gaaaccgcac caggacggcc aggacgaacc gtttttcatt accgaagaga 10981 tcgaggcgga gatgatcgcg gccgggtacg tgttcgagcc gcccgcgcac gtctcaaccg 11041 tgcggctgca tgaaatcctg gccggtttgt ctgatgccaa gctggcggcc tggccggcca 11101 gcttggccgc tgaagaaacc gagcgccgcc gtctaaaaag gtgatgtgta tttgagtaaa 11161 acagcttgcg tcatgcggtc gctgcgtata tgatgcgatg agtaaataaa caaatacgca 11221 aggggaacgc atgaaggtta tcgctgtact taaccagaaa ggcgggtcag gcaagacgac 11281 catcgcaacc catctagccc gcgccctgca actcgccggg gccgatgttc tgttagtcga 11341 ttccgatccc cagggcagtg cccgcgattg ggcggccgtg cgggaagatc aaccgctaac 11401 cgttgtcggc atcgaccgcc cgacgattga ccgcgacgtg aaggccatcg gccggcgcga 11461 cttcgtagtg atcgacggag cgccccaggc ggcggacttg gctgtgtccg cgatcaaggc 11521 agccgacttc gtgctgattc cggtgcagcc aagcccttac gacatatggg ccaccgccga 11581 cctggtggag ctggttaagc agcgcattga ggtcacggat ggaaggctac aagcggcctt 11641 tgtcgtgtcg cgggcgatca aaggcacgcg catcggcggt gaggttgccg aggcgctggc 11701 cgggtacgag ctgcccattc ttgagtcccg tatcacgcag cgcgtgagct acccaggcac 11761 tgccgccgcc ggcacaaccg ttcttgaatc agaacccgag ggcgacgctg cccgcgaggt 11821 ccaggcgctg gccgctgaaa ttaaatcaaa actcatttga gttaatgagg taaagagaaa 11881 atgagcaaaa gcacaaacac gctaagtgcc ggccgtccga gcgcacgcag cagcaaggct 11941 gcaacgttgg ccagcctggc agacacgcca gccatgaagc gggtcaactt tcagttgccg 12001 gcggaggatc acaccaagct gaagatgtac gcggtacgcc aaggcaagac cattaccgag 12061 ctgctatctg aatacatcgc gcagctacca gagtaaatga gcaaatgaat aaatgagtag 12121 atgaatttta gcggctaaag gaggcggcat ggaaaatcaa gaacaaccag gcaccgacgc 12181 cgtggaatgc cccatgtgtg gaggaacggg cggttggcca ggcgtaagcg gctgggttgc 12241 ctgccggccc tgcaatggca ctggaacccc caagcccgag gaatcggcgt gagcggtcgc 12301 aaaccatccg gcccggtaca aatcggcgcg gcgctgggtg atgacctggt ggagaagttg 12361 aaggccgcgc aggccgccca gcggcaacgc atcgaggcag aagcacgccc cggtgaatcg 12421 tggcaagcgg ccgctgatcg aatccgcaaa gaatcccggc aaccgccggc agccggtgcg 12481 ccgtcgatta ggaagccgcc caagggcgac gagcaaccag attttttcgt tccgatgctc 12541 tatgacgtgg gcacccgcga tagtcgcagc atcatggacg tggccgtttt ccgtctgtcg 12601 aagcgtgacc gacgagctgg cgaggtgatc cgctacgagc ttccagacgg gcacgtagag 12661 gtttccgcag ggccggccgg catggccagt gtgtgggatt acgacctggt actgatggcg 12721 gtttcccatc taaccgaatc catgaaccga taccgggaag ggaagggaga caagcccggc 12781 cgcgtgttcc gtccacacgt tgcggacgta ctcaagttct gccggcgagc cgatggcgga 12841 aagcagaaag acgacctggt agaaacctgc attcggttaa acaccacgca cgttgccatg 12901 cagcgtacga agaaggccaa gaacggccgc ctggtgacgg tatccgaggg tgaagccttg 12961 attagccgct acaagatcgt aaagagcgaa accgggcggc cggagtacat cgagatcgag 13021 ctagctgatt ggatgtaccg cgagatcaca gaaggcaaga acccggacgt gctgacggtt 13081 caccccgatt actttttgat cgatcccggc atcggccgtt ttctctaccg cctggcacgc 13141 cgcgccgcag gcaaggcaga agccagatgg ttgttcaaga cgatctacga acgcagtggc 13201 agcgccggag agttcaagaa gttctgtttc accgtgcgca agctgatcgg gtcaaatgac 13261 ctgccggagt acgatttgaa ggaggaggcg gggcaggctg gcccgatcct agtcatgcgc 13321 taccgcaacc tgatcgaggg cgaagcatcc gccggttcct aatgtacgga gcagatgcta 13381 gggcaaattg ccctagcagg ggaaaaaggt cgaaaaggtc tctttcctgt ggatagcacg 13441 tacattggga acccaaagcc gtacattggg aaccggaacc cgtacattgg gaacccaaag 13501 ccgtacattg ggaaccggtc acacatgtaa gtgactgata taaaagagaa aaaaggcgat 13561 ttttccgcct aaaactcttt aaaacttatt aaaactctta aaacccgcct ggcctgtgca 13621 taactgtctg gccagcgcac agcccaagag ctgcaaaaag cgcctaccct tcggtcgctg 13681 cgctccctac gccccgccgc ttcgcgtcgg cctatcgcgg ccgctggccg ctcaaaaatg 13741 gctggcctac ggccaggcaa tctaccaggg cgcggacaag ccgcgccgtc gccactcgac 13801 cgccggcgcc cacatcaagg caccctgcct cgcgcgtttc ggtgatgacg gtgaaaacct 13861 ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag 13921 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca 13981 gtcacgtagc gatagcggag tgtatactgg cttaactatg cggcatcaga gcagattgta 14041 ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc 14101 atcaggccct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg 14161 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac 14221 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg 14281 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca 14341 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc 14401 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc 14461 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag 14521 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc 14581 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca 14641 gcagccactg gtaacaggat tagcagagcg aggtat //
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