质粒名称: | pLentiCRISPR-E |
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出品公司: | 爱迪基因 |
目录编号: | 78852 |
存储实验室: | Phillip Abbosh |
载体骨架基本信息 |
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载体名称: | pLentiCRISPRV2 |
载体抗性: | 氨苄霉素 |
空载体大小: | 14849 bp |
载体修饰: | espcas9: K848A, K1003A, K1060A |
载体类型: | 哺乳细胞表达,Lentiviral, CRISPR |
筛选标记: | 嘌呤霉素 |
高拷贝/低拷贝: | 高拷贝 |
生长条件: | 37 ℃ |
插入基因信息 |
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标签/融合蛋白: | Cas9-P2A-Puro |
克隆方法: | 限制性内切酶 |
文献引用方法 |
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材料和方法部分: | pLentiCRISPR-E was a gift from Phillip Abbosh (爱迪基因 plasmid # 78852) |
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 pLentiCRISPR-E 14849 bp ds-DNA circular SYN 10-JAN-2018 DEFINITION Introduce sgRNA into a lentiviral vector (LentiCRISPR V2) which contains eSpCas9 and puromycin cassette. ACCESSION . VERSION . KEYWORDS pLentiCRISPR-E SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 14849) TITLE LentiCRISPR version E JOURNAL Unpublished COMMENT This file is created by Vector NTI http://www.biofeng.com/ COMMENT ORIGDB|GenBank COMMENT VNTAUTHORNAME|biofeng.com| COMMENT VNTNAME|pLentiCRISPR-E| FEATURES Location/Qualifiers source 1..14849 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(47..66) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 238..617 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 619..817 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 772..792 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" LTR 835..1015 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 1062..1187 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1680..1913 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." misc_feature 2440..2557 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 2608..2848 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 2608..2628 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 2779..2798 /label=LKO.1 5' /note="Human U6 promoter, forward primer" misc_RNA 4738..4813 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" promoter 4875..5086 /label=EF-1-alpha core promoter /note="core promoter for human elongation factor EF-1-alpha" regulatory 5105..5114 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 5111..9214 /codon_start=1 /product="Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system, mutated to improve targeting specificity (Slaymaker et al., 2016)" /label=eSpCas9(1.1) /note="carries the mutations K848A, K1003A, and R1060A" /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLADDSIDNKVLTRSDKNRGKSDNVP SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL NAVVGTALIKKYPALESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT EITLANGEIRKAPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL DATLIHQSITGLYETRIDLSQLGGD" CDS 9215..9262 /codon_start=1 /product="bipartite nuclear localization signal from nucleoplasmin" /label=nucleoplasmin NLS /translation="KRPAATKKAGQAKKKK" CDS 9272..9328 /codon_start=1 /product="2A peptide from porcine teschovirus-1 polyprotein" /label=P2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="ATNFSLLKQAGDVEENPGP" CDS 9329..9925 /codon_start=1 /gene="pac from Streptomyces alboniger" /product="puromycin N-acetyltransferase" /label=PuroR /note="confers resistance to puromycin" /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA" primer_bind 9822..9842 /label=Puro-F /note="Puromycin resistance gene, forward primer" misc_feature 9941..10529 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(9994..10014) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(10412..10423) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 10601..10834 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" primer_bind complement(10860..10877) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 10866..11090 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 11136..11564 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(11223..11242) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 11433..11454 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(11573..11593) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 11578..11907 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 11758..11893 /label=SV40 ori /note="SV40 origin of replication" primer_bind 11820..11839 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 11955..12002 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 12021..12395 /codon_start=1 /gene="Sh ble from Streptoalloteichus hindustanus" /product="antibiotic-binding protein" /label=BleoR /note="confers resistance to bleomycin, phleomycin, and Zeocin(TM)" /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR EFALRDPAGNCVHFVAEEQD" polyA_signal 12525..12646 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(12562..12581) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 12616..12635 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(12695..12711) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(12695..12711) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(12708..12730) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 12719..12735 /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(12743..12773) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 12788..12809 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(12926..12943) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(13097..13685) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(13177..13196) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(13856..14716) /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 14479..14498 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(14717..14821) /gene="bla" /label=AmpR promoter ORIGIN 1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg 61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt 121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc 181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac 241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat 301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg 361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt 421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag 481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc 541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag 601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt 661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc 721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg 781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct 841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt 901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac 961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc 1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc 1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa 1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg 1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata 1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc 1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga 1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc 1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca 1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg 1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga 1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata 1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg 1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg 1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag 1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt 1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt 1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt 2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag 2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata 2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta 2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta 2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa 2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt 2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat 2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa 2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag 2581 agatccagtt tggttaatta aggtaccgag ggcctatttc ccatgattcc ttcatatttg 2641 catatacgat acaaggctgt tagagagata attagaatta atttgactgt aaacacaaag 2701 atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta 2761 aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc 2821 ttggctttat atatcttgtg gaaaggacga aacaccggag acggttgtaa atgagcacac 2881 aaaatacaca tgctaaaata ttatattcta tgacctttat aaaatcaacc aaaatcttct 2941 ttttaataac tttagtatca ataattagaa tttttatgtt cctttttgca aacttttaat 3001 aaaaatgagc aaaataaaaa aacgctagtt ttagtaactc gcgttgtttt cttcaccttt 3061 aataatagct actccaccac ttgttcctaa gcggtcagct cctgcttcaa tcattttttg 3121 agcatcttca aatgttctaa ctccaccagc tgctttaact aaagcattgt ctttaacaac 3181 tgacttcatt agtttaacat cttcaaatgt tgcacctgat tttgaaaatc ctgttgatgt 3241 tttaacaaat tctaatccag cttcaacagc tatttcacaa gctttcatga tttcttcttt 3301 tgttaataaa caattttcca taatacattt aacaacatgt gatccagctg ctttttttac 3361 agctttcatg tcttctaaaa ctaattcata atttttgtct tttaatgcac caatatttaa 3421 taccatatca atttctgttg caccatcttt aattgcttca gaaacttcga atgcttttgt 3481 agctgttgtg catgcaccta gaggaaaacc tacaacattt gttattccta catttgtgcc 3541 ttttaataat tctttacaat agcttgttca atatgaatta acacaaactg ttgcaaaatc 3601 aaattcaatt gcttcatcac ataattgttt aatttcagct ttcgtagcat cttgttttaa 3661 taatgtgtga tctatatatt tgtttagttt cattttttct cctatatatt catttttaat 3721 tttaattctt taataatttc gtctacttta actttagcgt tttgaacaga ttcaccaaca 3781 cctataaaat aaatttttag tttaggttca gttccacttg ggcgaacagc aaatcatgac 3841 ttatcttcta aataaaattt tagtaagtct tgtcctggca tattatacat tccatcgatg 3901 tagtcttcaa cattaacaac tttaagtcca gcaatttgag ttaagggtgt tgctctcaat 3961 gatttcatta atggttcaat ttttaatttc ttttcttctg gtttaaaatt caagtttaaa 4021 gtgaaagtgt aatatgcacc catttcttta aataaatctt ctaaatagtc tactaatgtt 4081 ttattttgtt ttttataaaa tcaagcagcc tctgctatta atatagaagc ttgtattcca 4141 tctttatctc tagctgagtc atcaattaca tatccataac tttcttcata agcaaaaaca 4201 aaatttaatc cgttatcttc ttctttagca atttctctac ccattcattt aaatccagtt 4261 aaagttttta caatattaac tccatatttt tcatgagcga ttctatcacc caaatcactt 4321 gttacaaaac ttgaatatag agccggattt tttggaatgc tatttaagcg ttttagattt 4381 gataattttc aatcaattaa aattggtcct gtttgatttc catctaatct tacaaaatga 4441 ccatcatgtt ttattgccat tccaaatctg tcagcatctg ggtcattcat aataataata 4501 tctgcatcat gtttaatacc atattcaagc ggtatttttc atgcaggatc aaattctgga 4561 tttggattta caacattttt aaatgtttca tcttcaaatg catgctcttc aacctcaata 4621 acgttatatc ctgattcacg taatattttt ggggtaaatt tagttcctgt tccattaact 4681 gcgctaaaaa taatttttaa atctttttta gcttcttgct cttttttgta cgtctctgtt 4741 ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg aaaaagtggc 4801 accgagtcgg tgcttttttg aattcgctag ctaggtcttg aaaggagtgg gaattggctc 4861 cggtgcccgt cagtgggcag agcgcacatc gcccacagtc cccgagaagt tggggggagg 4921 ggtcggcaat tgatccggtg cctagagaag gtggcgcggg gtaaactggg aaagtgatgt 4981 cgtgtactgg ctccgccttt ttcccgaggg tgggggagaa ccgtatataa gtgcagtagt 5041 cgccgtgaac gttctttttc gcaacgggtt tgccgccaga acacaggacc ggttctagag 5101 cgctgccacc atggacaaga agtacagcat cggcctggac atcggcacca actctgtggg 5161 ctgggccgtg atcaccgacg agtacaaggt gcccagcaag aaattcaagg tgctgggcaa 5221 caccgaccgg cacagcatca agaagaacct gatcggagcc ctgctgttcg acagcggcga 5281 aacagccgag gccacccggc tgaagagaac cgccagaaga agatacacca gacggaagaa 5341 ccggatctgc tatctgcaag agatcttcag caacgagatg gccaaggtgg acgacagctt 5401 cttccacaga ctggaagagt ccttcctggt ggaagaggat aagaagcacg agcggcaccc 5461 catcttcggc aacatcgtgg acgaggtggc ctaccacgag aagtacccca ccatctacca 5521 cctgagaaag aaactggtgg acagcaccga caaggccgac ctgcggctga tctatctggc 5581 cctggcccac atgatcaagt tccggggcca cttcctgatc gagggcgacc tgaaccccga 5641 caacagcgac gtggacaagc tgttcatcca gctggtgcag acctacaacc agctgttcga 5701 ggaaaacccc atcaacgcca gcggcgtgga cgccaaggcc atcctgtctg ccagactgag 5761 caagagcaga cggctggaaa atctgatcgc ccagctgccc ggcgagaaga agaatggcct 5821 gttcggaaac ctgattgccc tgagcctggg cctgaccccc aacttcaaga gcaacttcga 5881 cctggccgag gatgccaaac tgcagctgag caaggacacc tacgacgacg acctggacaa 5941 cctgctggcc cagatcggcg accagtacgc cgacctgttt ctggccgcca agaacctgtc 6001 cgacgccatc ctgctgagcg acatcctgag agtgaacacc gagatcacca aggcccccct 6061 gagcgcctct atgatcaaga gatacgacga gcaccaccag gacctgaccc tgctgaaagc 6121 tctcgtgcgg cagcagctgc ctgagaagta caaagagatt ttcttcgacc agagcaagaa 6181 cggctacgcc ggctacattg acggcggagc cagccaggaa gagttctaca agttcatcaa 6241 gcccatcctg gaaaagatgg acggcaccga ggaactgctc gtgaagctga acagagagga 6301 cctgctgcgg aagcagcgga ccttcgacaa cggcagcatc ccccaccaga tccacctggg 6361 agagctgcac gccattctgc ggcggcagga agatttttac ccattcctga aggacaaccg 6421 ggaaaagatc gagaagatcc tgaccttccg catcccctac tacgtgggcc ctctggccag 6481 gggaaacagc agattcgcct ggatgaccag aaagagcgag gaaaccatca ccccctggaa 6541 cttcgaggaa gtggtggaca agggcgcttc cgcccagagc ttcatcgagc ggatgaccaa 6601 cttcgataag aacctgccca acgagaaggt gctgcccaag cacagcctgc tgtacgagta 6661 cttcaccgtg tataacgagc tgaccaaagt gaaatacgtg accgagggaa tgagaaagcc 6721 cgccttcctg agcggcgagc agaaaaaggc catcgtggac ctgctgttca agaccaaccg 6781 gaaagtgacc gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt gcttcgactc 6841 cgtggaaatc tccggcgtgg aagatcggtt caacgcctcc ctgggcacat accacgatct 6901 gctgaaaatt atcaaggaca aggacttcct ggacaatgag gaaaacgagg acattctgga 6961 agatatcgtg ctgaccctga cactgtttga ggacagagag atgatcgagg aacggctgaa 7021 aacctatgcc cacctgttcg acgacaaagt gatgaagcag ctgaagcggc ggagatacac 7081 cggctggggc aggctgagcc ggaagctgat caacggcatc cgggacaagc agtccggcaa 7141 gacaatcctg gatttcctga agtccgacgg cttcgccaac agaaacttca tgcagctgat 7201 ccacgacgac agcctgacct ttaaagagga catccagaaa gcccaggtgt ccggccaggg 7261 cgatagcctg cacgagcaca ttgccaatct ggccggcagc cccgccatta agaagggcat 7321 cctgcagaca gtgaaggtgg tggacgagct cgtgaaagtg atgggccggc acaagcccga 7381 gaacatcgtg atcgaaatgg ccagagagaa ccagaccacc cagaagggac agaagaacag 7441 ccgcgagaga atgaagcgga tcgaagaggg catcaaagag ctgggcagcc agatcctgaa 7501 agaacacccc gtggaaaaca cccagctgca gaacgagaag ctgtacctgt actacctgca 7561 gaatgggcgg gatatgtacg tggaccagga actggacatc aaccggctgt ccgactacga 7621 tgtggaccat atcgtgcctc agagctttct ggccgacgac tccatcgaca acaaggtgct 7681 gaccagaagc gacaagaacc ggggcaagag cgacaacgtg ccctccgaag aggtcgtgaa 7741 gaagatgaag aactactggc ggcagctgct gaacgccaag ctgattaccc agagaaagtt 7801 cgacaatctg accaaggccg agagaggcgg cctgagcgaa ctggataagg ccggcttcat 7861 caagagacag ctggtggaaa cccggcagat cacaaagcac gtggcacaga tcctggactc 7921 ccggatgaac actaagtacg acgagaatga caagctgatc cgggaagtga aagtgatcac 7981 cctgaagtcc aagctggtgt ccgatttccg gaaggatttc cagttttaca aagtgcgcga 8041 gatcaacaac taccaccacg cccacgacgc ctacctgaac gccgtcgtgg gaaccgccct 8101 gatcaaaaag taccctgcgc tggaaagcga gttcgtgtac ggcgactaca aggtgtacga 8161 cgtgcggaag atgatcgcca agagcgagca ggaaatcggc aaggctaccg ccaagtactt 8221 cttctacagc aacatcatga actttttcaa gaccgagatt accctggcca acggcgagat 8281 ccggaaggcg cctctgatcg agacaaacgg cgaaaccggg gagatcgtgt gggataaggg 8341 ccgggatttt gccaccgtgc ggaaagtgct gagcatgccc caagtgaata tcgtgaaaaa 8401 gaccgaggtg cagacaggcg gcttcagcaa agagtctatc ctgcccaaga ggaacagcga 8461 taagctgatc gccagaaaga aggactggga ccctaagaag tacggcggct tcgacagccc 8521 caccgtggcc tattctgtgc tggtggtggc caaagtggaa aagggcaagt ccaagaaact 8581 gaagagtgtg aaagagctgc tggggatcac catcatggaa agaagcagct tcgagaagaa 8641 tcccatcgac tttctggaag ccaagggcta caaagaagtg aaaaaggacc tgatcatcaa 8701 gctgcctaag tactccctgt tcgagctgga aaacggccgg aagagaatgc tggcctctgc 8761 cggcgaactg cagaagggaa acgaactggc cctgccctcc aaatatgtga acttcctgta 8821 cctggccagc cactatgaga agctgaaggg ctcccccgag gataatgagc agaaacagct 8881 gtttgtggaa cagcacaagc actacctgga cgagatcatc gagcagatca gcgagttctc 8941 caagagagtg atcctggccg acgctaatct ggacaaagtg ctgtccgcct acaacaagca 9001 ccgggataag cccatcagag agcaggccga gaatatcatc cacctgttta ccctgaccaa 9061 tctgggagcc cctgccgcct tcaagtactt tgacaccacc atcgaccgga agaggtacac 9121 cagcaccaaa gaggtgctgg acgccaccct gatccaccag agcatcaccg gcctgtacga 9181 gacacggatc gacctgtctc agctgggagg cgacaaaagg ccggcggcca cgaaaaaggc 9241 cggccaggca aaaaagaaaa agggatccgg cgcaacaaac ttctctctgc tgaaacaagc 9301 cggagatgtc gaagagaatc ctggaccgac cgagtacaag cccacggtgc gcctcgccac 9361 ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc 9421 cacgcgccac accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact 9481 cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc 9541 ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg 9601 cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct 9661 cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg gagtctcgcc 9721 cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga 9781 gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga 9841 gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg 9901 catgacccgc aagcccggtg cctgaacgcg ttaagtcgac aatcaacctc tggattacaa 9961 aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata 10021 cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc 10081 cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg 10141 tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac 10201 ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat 10261 cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt 10321 ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg ccacctggat 10381 tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc 10441 ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag 10501 tcggatctcc ctttgggccg cctccccgcg tcgactttaa gaccaatgac ttacaaggca 10561 gctgtagatc ttagccactt tttaaaagaa aaggggggac tggaagggct aattcactcc 10621 caacgaagac aagatctgct ttttgcttgt actgggtctc tctggttaga ccagatctga 10681 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct 10741 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc 10801 agaccctttt agtcagtgtg gaaaatctct agcagggccc gtttaaaccc gctgatcagc 10861 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt 10921 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca 10981 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga 11041 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc 11101 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag 11161 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc 11221 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc 11281 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa 11341 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg 11401 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac 11461 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta 11521 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg 11581 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg 11641 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt 11701 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc 11761 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt 11821 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc 11881 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga 11941 tctgatcagc acgtgttgac aattaatcat cggcatagta tatcggcata gtataatacg 12001 acaaggtgag gaactaaacc atggccaagt tgaccagtgc cgttccggtg ctcaccgcgc 12061 gcgacgtcgc cggagcggtc gagttctgga ccgaccggct cgggttctcc cgggacttcg 12121 tggaggacga cttcgccggt gtggtccggg acgacgtgac cctgttcatc agcgcggtcc 12181 aggaccaggt ggtgccggac aacaccctgg cctgggtgtg ggtgcgcggc ctggacgagc 12241 tgtacgccga gtggtcggag gtcgtgtcca cgaacttccg ggacgcctcc gggccggcca 12301 tgaccgagat cggcgagcag ccgtgggggc gggagttcgc cctgcgcgac ccggccggca 12361 actgcgtgca cttcgtggcc gaggagcagg actgacacgt gctacgagat ttcgattcca 12421 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga 12481 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag 12541 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt 12601 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac 12661 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt 12721 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg 12781 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt 12841 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt 12901 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc 12961 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg 13021 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg 13081 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac 13141 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg 13201 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct 13261 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg 13321 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct 13381 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac 13441 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt 13501 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc 13561 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca 13621 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat 13681 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac 13741 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt 13801 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc 13861 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg 13921 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg 13981 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc 14041 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta 14101 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg 14161 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct 14221 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta 14281 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg 14341 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga 14401 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt 14461 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca 14521 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt 14581 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt 14641 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga 14701 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt 14761 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc 14821 gcacatttcc ccgaaaagtg ccacctgac //