| 质粒名称: | eSpCas9(1.1)-Puro |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 117686 |
| 存储实验室: | Luc Leyns |
|
载体骨架基本信息 |
|
| 载体名称: | pSpCas9(BB)-2A-Puro |
| 载体出品公司: | Feng Zhang Lab |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 9175 bp |
| 完整质粒大小: | 9175 bp |
| 载体修饰: | K (AAG) 848 A (GCC), K (AAG) 1003 A (GCG), R (CGG) 1060 A (GCG) |
| 载体类型: | 哺乳细胞表达,CRISPR |
| 筛选标记: | 嘌呤霉素 |
| 高拷贝/低拷贝: | 未知 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | eSpCas9(1.1),K848A, K1003A, R1060A |
| 物种来源: | 合成 |
| 插入大小: | -- |
| 基因突变: | K848A, K1003A, R1060A |
| GenBank编号 | -- |
| 启动子 | U6 |
| 标签/融合蛋白: | -- |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | ApaI (未破坏) |
| 3' 克隆位点: | BsmI (未破坏) |
| 5' 测序引物 | ACCCATTCCTGAAGGACAAC |
| 3' 测序引物 | TGGCCAGGTACAGGAAGTTC |
|
文献引用方法 |
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| 材料和方法部分: | eSpCas9(1.1)-Puro was a gift from Luc Leyns (Addgene plasmid # 117686 ; http://n2t.net/addgene:117686 ; RRID:Addgene_117686) |
| 参考文献部分: | Loss of Emp2 compromises cardiogenic differentiation in mouse embryonic stem cells. Liu Y, Dakou E, Meng Y, Leyns L. Biochem Biophys Res Commun. 2019 Feb 14. pii: S0006-291X(19)30235-9. doi: 10.1016/j.bbrc.2019.02.048. 10.1016/j.bbrc.2019.02.048 PubMed 30773261 |
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
-Puro质粒图谱.png)
载体简介
载体序列
LOCUS eSpCas9(1.1)-Puro 9034 bp ds-DNA circular SYN 22-FEB-2019 DEFINITION The plasmids pSpCas9 (BB)-2A-Puro (Addgene, #48139) and eSpCas9 (1.1) (Addgene, #71814) were combined to express both "enhanced specificity" Cas9 and puromycin resistance protein. . ACCESSION . VERSION . KEYWORDS eSpCas9(1.1)-Puro SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 9034) TITLE eSpCas9(1.1) + Puromycin selection JOURNAL Unpublished REFERENCE 2 (bases 1 to 9034) AUTHORS . TITLE Direct Submission JOURNAL Exported Feb 22, 2019 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..9034 /organism="synthetic DNA construct" /mol_type="other DNA" intron 27..254 /label=hybrid intron /note="hybrid between chicken beta-actin (CBA) and minute virus of mice (MMV) introns (Gray et al., 2011)" regulatory 266..275 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 275..340 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" CDS 347..367 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 392..4492 /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLADDSIDNKVLTRSDKNRGKSDNVPS EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN AVVGTALIKKYPALESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE ITLANGEIRKAPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD ATLIHQSITGLYETRIDLSQLGGD" CDS 4493..4540 /codon_start=1 /product="bipartite nuclear localization signal from nucleoplasmin" /label=nucleoplasmin NLS /translation="KRPAATKKAGQAKKKK" CDS 4556..4609 /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 4610..5209 /codon_start=1 /gene="pac from Streptomyces alboniger" /product="puromycin N-acetyltransferase" /label=PuroR /note="confers resistance to puromycin" /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS APHNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA" primer_bind complement(4610..4629) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind complement(5237..5254) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 5243..5450 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" repeat_region 5459..5599 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 5459..5588 /label=AAV2 ITR rep_origin 5674..6129 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(5761..5780) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 5971..5992 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(6146..6165) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 6265..6287 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(6325..6343) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 6411..6515 /gene="bla" /label=AmpR promoter CDS 6516..7376 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind complement(6734..6753) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 7547..8135 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 8036..8055 /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 8197..8437 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 8197..8217 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 8368..8387 /label=LKO.1 5' /note="Human U6 promoter, forward primer" misc_RNA 8464..8539 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" enhancer 8636..8921 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer; contains an 18-bp deletion relative to the standard CMV enhancer" ORIGIN 1 ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg ctgccttcgc cccgtgcccc 61 gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta ctcccacagg 121 tgagcgggcg ggacggccct tctcctccgg gctgtaatta gctgagcaag aggtaagggt 181 ttaagggatg gttggttggt ggggtattaa tgtttaatta cctggagcac ctgcctgaaa 241 tcactttttt tcaggttgga ccggtgccac catggactat aaggaccacg acggagacta 301 caaggatcat gatattgatt acaaagacga tgacgataag atggccccaa agaagaagcg 361 gaaggtcggt atccacggag tcccagcagc cgacaagaag tacagcatcg gcctggacat 421 cggcaccaac tctgtgggct gggccgtgat caccgacgag tacaaggtgc ccagcaagaa 481 attcaaggtg ctgggcaaca ccgaccggca cagcatcaag aagaacctga tcggagccct 541 gctgttcgac agcggcgaaa cagccgaggc cacccggctg aagagaaccg ccagaagaag 601 atacaccaga cggaagaacc ggatctgcta tctgcaagag atcttcagca acgagatggc 661 caaggtggac gacagcttct tccacagact ggaagagtcc ttcctggtgg aagaggataa 721 gaagcacgag cggcacccca tcttcggcaa catcgtggac gaggtggcct accacgagaa 781 gtaccccacc atctaccacc tgagaaagaa actggtggac agcaccgaca aggccgacct 841 gcggctgatc tatctggccc tggcccacat gatcaagttc cggggccact tcctgatcga 901 gggcgacctg aaccccgaca acagcgacgt ggacaagctg ttcatccagc tggtgcagac 961 ctacaaccag ctgttcgagg aaaaccccat caacgccagc ggcgtggacg ccaaggccat 1021 cctgtctgcc agactgagca agagcagacg gctggaaaat ctgatcgccc agctgcccgg 1081 cgagaagaag aatggcctgt tcggaaacct gattgccctg agcctgggcc tgacccccaa 1141 cttcaagagc aacttcgacc tggccgagga tgccaaactg cagctgagca aggacaccta 1201 cgacgacgac ctggacaacc tgctggccca gatcggcgac cagtacgccg acctgtttct 1261 ggccgccaag aacctgtccg acgccatcct gctgagcgac atcctgagag tgaacaccga 1321 gatcaccaag gcccccctga gcgcctctat gatcaagaga tacgacgagc accaccagga 1381 cctgaccctg ctgaaagctc tcgtgcggca gcagctgcct gagaagtaca aagagatttt 1441 cttcgaccag agcaagaacg gctacgccgg ctacattgac ggcggagcca gccaggaaga 1501 gttctacaag ttcatcaagc ccatcctgga aaagatggac ggcaccgagg aactgctcgt 1561 gaagctgaac agagaggacc tgctgcggaa gcagcggacc ttcgacaacg gcagcatccc 1621 ccaccagatc cacctgggag agctgcacgc cattctgcgg cggcaggaag atttttaccc 1681 attcctgaag gacaaccggg aaaagatcga gaagatcctg accttccgca tcccctacta 1741 cgtgggccct ctggccaggg gaaacagcag attcgcctgg atgaccagaa agagcgagga 1801 aaccatcacc ccctggaact tcgaggaagt ggtggacaag ggcgcttccg cccagagctt 1861 catcgagcgg atgaccaact tcgataagaa cctgcccaac gagaaggtgc tgcccaagca 1921 cagcctgctg tacgagtact tcaccgtgta taacgagctg accaaagtga aatacgtgac 1981 cgagggaatg agaaagcccg ccttcctgag cggcgagcag aaaaaggcca tcgtggacct 2041 gctgttcaag accaaccgga aagtgaccgt gaagcagctg aaagaggact acttcaagaa 2101 aatcgagtgc ttcgactccg tggaaatctc cggcgtggaa gatcggttca acgcctccct 2161 gggcacatac cacgatctgc tgaaaattat caaggacaag gacttcctgg acaatgagga 2221 aaacgaggac attctggaag atatcgtgct gaccctgaca ctgtttgagg acagagagat 2281 gatcgaggaa cggctgaaaa cctatgccca cctgttcgac gacaaagtga tgaagcagct 2341 gaagcggcgg agatacaccg gctggggcag gctgagccgg aagctgatca acggcatccg 2401 ggacaagcag tccggcaaga caatcctgga tttcctgaag tccgacggct tcgccaacag 2461 aaacttcatg cagctgatcc acgacgacag cctgaccttt aaagaggaca tccagaaagc 2521 ccaggtgtcc ggccagggcg atagcctgca cgagcacatt gccaatctgg ccggcagccc 2581 cgccattaag aagggcatcc tgcagacagt gaaggtggtg gacgagctcg tgaaagtgat 2641 gggccggcac aagcccgaga acatcgtgat cgaaatggcc agagagaacc agaccaccca 2701 gaagggacag aagaacagcc gcgagagaat gaagcggatc gaagagggca tcaaagagct 2761 gggcagccag atcctgaaag aacaccccgt ggaaaacacc cagctgcaga acgagaagct 2821 gtacctgtac tacctgcaga atgggcggga tatgtacgtg gaccaggaac tggacatcaa 2881 ccggctgtcc gactacgatg tggaccatat cgtgcctcag agctttctgg ccgacgactc 2941 catcgacaac aaggtgctga ccagaagcga caagaaccgg ggcaagagcg acaacgtgcc 3001 ctccgaagag gtcgtgaaga agatgaagaa ctactggcgg cagctgctga acgccaagct 3061 gattacccag agaaagttcg acaatctgac caaggccgag agaggcggcc tgagcgaact 3121 ggataaggcc ggcttcatca agagacagct ggtggaaacc cggcagatca caaagcacgt 3181 ggcacagatc ctggactccc ggatgaacac taagtacgac gagaatgaca agctgatccg 3241 ggaagtgaaa gtgatcaccc tgaagtccaa gctggtgtcc gatttccgga aggatttcca 3301 gttttacaaa gtgcgcgaga tcaacaacta ccaccacgcc cacgacgcct acctgaacgc 3361 cgtcgtggga accgccctga tcaaaaagta ccctgcgctg gaaagcgagt tcgtgtacgg 3421 cgactacaag gtgtacgacg tgcggaagat gatcgccaag agcgagcagg aaatcggcaa 3481 ggctaccgcc aagtacttct tctacagcaa catcatgaac tttttcaaga ccgagattac 3541 cctggccaac ggcgagatcc ggaaggcgcc tctgatcgag acaaacggcg aaaccgggga 3601 gatcgtgtgg gataagggcc gggattttgc caccgtgcgg aaagtgctga gcatgcccca 3661 agtgaatatc gtgaaaaaga ccgaggtgca gacaggcggc ttcagcaaag agtctatcct 3721 gcccaagagg aacagcgata agctgatcgc cagaaagaag gactgggacc ctaagaagta 3781 cggcggcttc gacagcccca ccgtggccta ttctgtgctg gtggtggcca aagtggaaaa 3841 gggcaagtcc aagaaactga agagtgtgaa agagctgctg gggatcacca tcatggaaag 3901 aagcagcttc gagaagaatc ccatcgactt tctggaagcc aagggctaca aagaagtgaa 3961 aaaggacctg atcatcaagc tgcctaagta ctccctgttc gagctggaaa acggccggaa 4021 gagaatgctg gcctctgccg gcgaactgca gaagggaaac gaactggccc tgccctccaa 4081 atatgtgaac ttcctgtacc tggccagcca ctatgagaag ctgaagggct cccccgagga 4141 taatgagcag aaacagctgt ttgtggaaca gcacaagcac tacctggacg agatcatcga 4201 gcagatcagc gagttctcca agagagtgat cctggccgac gctaatctgg acaaagtgct 4261 gtccgcctac aacaagcacc gggataagcc catcagagag caggccgaga atatcatcca 4321 cctgtttacc ctgaccaatc tgggagcccc tgccgccttc aagtactttg acaccaccat 4381 cgaccggaag aggtacacca gcaccaaaga ggtgctggac gccaccctga tccaccagag 4441 catcaccggc ctgtacgaga cacggatcga cctgtctcag ctgggaggcg acaaaaggcc 4501 ggcggccacg aaaaaggccg gccaggcaaa aaagaaaaag gaattcggca gtggagaggg 4561 cagaggaagt ctgctaacat gcggtgacgt cgaggagaat cctggcccaa tgaccgagta 4621 caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc 4681 cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga 4741 gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt 4801 gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc 4861 gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc 4921 cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt 4981 cctggccacc gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt 5041 gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc 5101 gccccacaac ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt 5161 gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgag aattctaact 5221 agagctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc 5281 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat 5341 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg 5401 caggacagca agggggagga ttgggaagag aatagcaggc atgctgggga gcggccgcag 5461 gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc 5521 gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga 5581 gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca tctgtgcggt 5641 atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg 5701 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ttagcgcccg 5761 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc 5821 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa 5881 aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc 5941 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac 6001 tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt tcggtctatt 6061 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt 6121 ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc 6181 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg 6241 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat 6301 caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca 6361 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc 6421 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct 6481 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg 6541 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg 6601 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc 6661 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca 6721 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac 6781 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa 6841 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg 6901 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt 6961 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg 7021 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc 7081 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga 7141 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta 7201 ttgctgataa atctggagcc ggtgagcgtg gaagccgcgg tatcattgca gcactggggc 7261 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg 7321 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt 7381 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa 7441 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt 7501 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt 7561 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt 7621 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga 7681 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag 7741 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata 7801 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg 7861 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga 7921 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca 7981 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa 8041 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt 8101 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac 8161 ggttcctggc cttttgctgg ccttttgctc acatgtgagg gcctatttcc catgattcct 8221 tcatatttgc atatacgata caaggctgtt agagagataa ttggaattaa tttgactgta 8281 aacacaaaga tattagtaca aaatacgtga cgtagaaagt aataatttct tgggtagttt 8341 gcagttttaa aattatgttt taaaatggac tatcatatgc ttaccgtaac ttgaaagtat 8401 ttcgatttct tggctttata tatcttgtgg aaaggacgaa acaccgggtc ttcgagaaga 8461 cctgttttag agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa 8521 agtggcaccg agtcggtgct tttttgtttt agagctagaa atagcaagtt aaaataaggc 8581 tagtccgttt ttagcgcgtg cgccaattct gcagacaaat ggctctagag gtacccgtta 8641 cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt 8701 caatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa 8761 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca 8821 atgacggtaa atggcccgcc tggcattgtg cccagtacat gaccttatgg gactttccta 8881 cttggcagta catctacgta ttagtcatcg ctattaccat ggtcgaggtg agccccacgt 8941 tctgcttcac tctccccatc tcccccccct ccccaccccc aattttgtat ttatttattt 9001 tttaattatt ttgtgcagcg atgggggcgg gggc //
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