- 规格:20μl质粒
- 原核抗性:壮观霉素Spe
- 克隆菌株:大肠杆菌Stbl3
- 质粒简介: pX330S-3是一个哺乳细胞CAS9和gRNA表达质粒。
- 质粒图谱1: png/1-1G205092U3K2.png
质粒序列: 点击查看序列LOCUS Exported 8698 bp ds-DNA circular SYN 04-DEC-2017DEFINITION synthetic circular DNAFEATURES Location/Qualifierssource 1..8698/organism="synthetic DNA construct"/mol_type="other DNA"enhancer 8..293/label=CMV enhancer/note="human cytomegalovirus immediate early enhancer;contains an 18-bp deletion relative to the standard CMVenhancer"promoter 295..572/label=chicken beta-actin promoterintron 573..801/label=hybrid intron/note="hybrid between chicken beta-actin (CBA) and minutevirus of mice (MMV) introns (Gray et al., 2011)"regulatory 813..822/regulatory_class="other"/note="vertebrate consensus sequence for strong initiationof translation (Kozak, 1987)"CDS 822..887/codon_start=1/product="three tandem FLAG(R) epitope tags, followed by anenterokinase cleavage site"/label=3xFLAG/translation="DYKDHDGDYKDHDIDYKDDDDK"CDS 894..914/codon_start=1/product="nuclear localization signal of SV40 (simian virus40) large T antigen"/label=SV40 NLS/translation="PKKKRKV"CDS 939..5039/codon_start=1/product="Cas9 (Csn1) endonuclease from the Streptococcuspyogenes Type II CRISPR/Cas system"/label=Cas9/note="generates RNA-guided double strand breaks in DNA"/translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIHQSITGLYETRIDLSQLGGD"CDS 5040..5087/codon_start=1/product="bipartite nuclear localization signal fromnucleoplasmin"/label=nucleoplasmin NLS/translation="KRPAATKKAGQAKKKK"polyA_signal 5121..5328/label=bGH poly(A) signal/note="bovine growth hormone polyadenylation signal"repeat_region 5337..5477/label=AAV2 ITR/note="inverted terminal repeat of adeno-associated virusserotype 2"rep_origin 5552..6007/direction=RIGHT/label=f1 ori/note="f1 bacteriophage origin of replication; arrowindicates direction of (+) strand synthesis"CDS 6628..7419/codon_start=1/gene="aadA"/product="aminoglycoside adenylyltransferase (Murphy,1985)"/label=Spe/note="confers resistance to spectinomycin andstreptomycin"/translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPHSDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAKRELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLFEALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQPVILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"rep_origin 7602..8190/direction=RIGHT/label=ori/note="high-copy-number ColE1/pMB1/pBR322/pUC origin ofreplication"promoter 8267..8507/label=U6 promoter/note="RNA polymerase III promoter for human U6 snRNA"misc_RNA 8534..8609/label=gRNA scaffold/note="guide RNA scaffold for the Streptococcus pyogenesCRISPR/Cas9 system"ORIGIN1 tgagacccgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc61 gcccattgac gtcaatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt121 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc181 ctattgacgt caatgacggt aaatggcccg cctggcattg tgcccagtac atgaccttat241 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtcgagg301 tgagccccac gttctgcttc actctcccca tctccccccc ctccccaccc ccaattttgt361 atttatttat tttttaatta ttttgtgcag cgatgggggc gggggggggg ggggggcgcg421 cgccaggcgg ggcggggcgg ggcgaggggc ggggcggggc gaggcggaga ggtgcggcgg481 cagccaatca gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc541 ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc tgcgacgctg ccttcgcccc601 gtgccccgct ccgccgccgc ctcgcgccgc ccgccccggc tctgactgac cgcgttactc661 ccacaggtga gcgggcggga cggcccttct cctccgggct gtaattagct gagcaagagg721 taagggttta agggatggtt ggttggtggg gtattaatgt ttaattacct ggagcacctg781 cctgaaatca ctttttttca ggttggaccg gtgccaccat ggactataag gaccacgacg841 gagactacaa ggatcatgat attgattaca aagacgatga cgataagatg gccccaaaga901 agaagcggaa ggtcggtatc cacggagtcc cagcagccga caagaagtac agcatcggcc961 tggacatcgg caccaactct gtgggctggg ccgtgatcac cgacgagtac aaggtgccca1021 gcaagaaatt caaggtgctg ggcaacaccg accggcacag catcaagaag aacctgatcg1081 gagccctgct gttcgacagc ggcgaaacag ccgaggccac ccggctgaag agaaccgcca1141 gaagaagata caccagacgg aagaaccgga tctgctatct gcaagagatc ttcagcaacg1201 agatggccaa ggtggacgac agcttcttcc acagactgga agagtccttc ctggtggaag1261 aggataagaa gcacgagcgg caccccatct tcggcaacat cgtggacgag gtggcctacc1321 acgagaagta ccccaccatc taccacctga gaaagaaact ggtggacagc accgacaagg1381 ccgacctgcg gctgatctat ctggccctgg cccacatgat caagttccgg ggccacttcc1441 tgatcgaggg cgacctgaac cccgacaaca gcgacgtgga caagctgttc atccagctgg1501 tgcagaccta caaccagctg ttcgaggaaa accccatcaa cgccagcggc gtggacgcca1561 aggccatcct gtctgccaga ctgagcaaga gcagacggct ggaaaatctg atcgcccagc1621 tgcccggcga gaagaagaat ggcctgttcg gaaacctgat tgccctgagc ctgggcctga1681 cccccaactt caagagcaac ttcgacctgg ccgaggatgc caaactgcag ctgagcaagg1741 acacctacga cgacgacctg gacaacctgc tggcccagat cggcgaccag tacgccgacc1801 tgtttctggc cgccaagaac ctgtccgacg ccatcctgct gagcgacatc ctgagagtga1861 acaccgagat caccaaggcc cccctgagcg cctctatgat caagagatac gacgagcacc1921 accaggacct gaccctgctg aaagctctcg tgcggcagca gctgcctgag aagtacaaag1981 agattttctt cgaccagagc aagaacggct acgccggcta cattgacggc ggagccagcc2041 aggaagagtt ctacaagttc atcaagccca tcctggaaaa gatggacggc accgaggaac2101 tgctcgtgaa gctgaacaga gaggacctgc tgcggaagca gcggaccttc gacaacggca2161 gcatccccca ccagatccac ctgggagagc tgcacgccat tctgcggcgg caggaagatt2221 tttacccatt cctgaaggac aaccgggaaa agatcgagaa gatcctgacc ttccgcatcc2281 cctactacgt gggccctctg gccaggggaa acagcagatt cgcctggatg accagaaaga2341 gcgaggaaac catcaccccc tggaacttcg aggaagtggt ggacaagggc gcttccgccc2401 agagcttcat cgagcggatg accaacttcg ataagaacct gcccaacgag aaggtgctgc2461 ccaagcacag cctgctgtac gagtacttca ccgtgtataa cgagctgacc aaagtgaaat