质粒名称: | LentiRCas9-Lambda2 |
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出品公司: | Addgene |
目录编号: | 104184 |
存储实验室: | Eugene Yeo |
载体骨架基本信息 |
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载体名称: | LentiCRISPR v2 |
载体出品公司: | -- |
载体抗性: | 氨苄青霉素 |
空载体大小: | -- |
完整质粒大小: | -- |
载体修饰: | -- |
载体类型: | 慢病毒,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: | U6-sgRNA, EFS-PIN-dCas9 |
物种来源: | -- |
插入大小: | -- |
基因突变: | -- |
GenBank编号 | -- |
启动子 | -- |
标签/融合蛋白: | -- |
克隆方法: | Gibson克隆 |
5' 克隆位点: | -- |
3' 克隆位点: | -- |
5' 测序引物 | LKO1.5 |
3' 测序引物 | T3 |
文献引用方法 |
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材料和方法部分: | LentiRCas9-Lambda2 was a gift from Eugene Yeo (Addgene plasmid # 104184 ; http://n2t.net/addgene:104184 ; RRID:Addgene_104184) |
参考文献部分: | Elimination of Toxic Microsatellite Repeat Expansion RNA by RNA-Targeting Cas9. Batra R, Nelles DA, Pirie E, Blue SM, Marina RJ, Wang H, Chaim IA, Thomas JD, Zhang N, Nguyen V, Aigner S, Markmiller S, Xia G, Corbett KD, Swanson MS, Yeo GW. Cell. 2017 Aug 24;170(5):899-912.e10. doi: 10.1016/j.cell.2017.07.010. Epub 2017 Aug 10. 10.1016/j.cell.2017.07.010 PubMed 28803727 |
买家导航
LentiRCas9Lambda2载体基本信息
LentiRCas9Lambda2载体图谱质粒图谱和多克隆位点信息
LentiRCas9Lambda2载体简介
LentiRCas9Lambda2载体序列
LOCUS LentiRCas9-Lambda2 13752 bp ds-DNA circular SYN 06-AUG-2018 DEFINITION Lentiviral transfer vector that carries U6-driven non-targeting sgRNA using a modified scaffold (Chen et al. Cell 2013) and CMV-driven PIN-dCas9. Derived from LentiCRISPR v2 (Zhang lab). ACCESSION . VERSION . KEYWORDS LentiRCas9-Lambda2 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 13752) AUTHORS Batra R, Nelles DA, Pirie E, Blue SM, Marina RJ, Wang H, Chaim IA, Thomas JD, Zhang N, Nguyen V, Aigner S, Markmiller S, Xia G, Corbett KD, Swanson MS, Yeo GW TITLE Elimination of Toxic Microsatellite Repeat Expansion RNA by RNA-Targeting Cas9. JOURNAL Cell. 2017 Aug 24;170(5):899-912.e10. doi: 10.1016/j.cell.2017.07.010. Epub 2017 Aug 10. PUBMED 28803727 REFERENCE 2 (bases 1 to 13752) AUTHORS . TITLE Direct Submission JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..13752 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 1..380 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 382..580 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 535..555 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" LTR 598..778 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 825..950 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1443..1676 /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 2203..2320 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 2371..2611 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 2371..2391 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 2542..2561 /label=LKO.1 5' /note="Human U6 promoter, forward primer" primer_bind complement(2748..2764) /label=SK primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2748..2764) /label=pBluescriptSK /note="For pBluescript vector" primer_bind complement(2801..2821) /label=T3 /note="T3 promoter, forward primer" promoter complement(2801..2819) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" promoter 2881..3092 /label=EF-1-alpha core promoter /note="core promoter for human elongation factor EF-1-alpha" CDS 3720..7820 /codon_start=1 /product="catalytically dead mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system" /label=dCas9 /note="RNA-guided DNA-binding protein that lacks endonuclease activity due to the D10A mutation in the RuvC catalytic domain and the H840A mutation in the HNH catalytic domain" /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD ATLIHQSITGLYETRIDLSQLGGD" CDS 7824..7850 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" CDS 7869..7889 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 7896..7916 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 7938..7994 /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 7995..8591 /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 8488..8508 /label=Puro-F /note="Puromycin resistance gene, forward primer" misc_feature 8607..9195 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(8660..8680) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(9078..9089) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 9267..9500 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" primer_bind complement(9526..9543) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 9532..9756 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 9802..10230 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(9889..9908) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 10099..10120 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(10239..10259) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 10244..10573 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 10424..10559 /label=SV40 ori /note="SV40 origin of replication" primer_bind 10486..10505 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 10621..10668 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 10687..11061 /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 11191..11312 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(11228..11247) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 11282..11301 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(11361..11377) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(11361..11377) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(11374..11396) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 11385..11401 /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(11409..11439) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 11454..11475 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(11592..11609) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(11763..12351) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(11843..11862) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(12522..13382) /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 13145..13164 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(13383..13487) /gene="bla" /label=AmpR promoter primer_bind complement(13562..13581) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" ORIGIN 1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc 61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca 121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga 181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc 241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag cgcgttttgc ctgtactggg 601 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg 661 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt 721 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt 781 ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga 841 ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa 901 aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc 961 gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat 1021 ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg 1081 gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc 1141 agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc 1201 atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa 1261 acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct tcagacctgg aggaggagat 1321 atgagggaca attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta 1381 ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga 1441 ataggagctt tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca 1501 atgacgctga cggtacaggc cagacaatta ttgtctggta tagtgcagca gcagaacaat 1561 ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac tcacagtctg gggcatcaag 1621 cagctccagg caagaatcct ggctgtggaa agatacctaa aggatcaaca gctcctgggg 1681 atttggggtt gctctggaaa actcatttgc accactgctg tgccttggaa tgctagttgg 1741 agtaataaat ctctggaaca gatttggaat cacacgacct ggatggagtg ggacagagaa 1801 attaacaatt acacaagctt aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa 1861 aagaatgaac aagaattatt ggaattagat aaatgggcaa gtttgtggaa ttggtttaac 1921 ataacaaatt ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt 1981 ttaagaatag tttttgctgt actttctata gtgaatagag ttaggcaggg atattcacca 2041 ttatcgtttc agacccacct cccaaccccg aggggacccg acaggcccga aggaatagaa 2101 gaagaaggtg gagagagaga cagagacaga tccattcgat tagtgaacgg atcggcactg 2161 cgtgcgccaa ttctgcagac aaatggcagt attcatccac aattttaaaa gaaaaggggg 2221 gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac 2281 taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag 2341 cagagatcca gtttggttaa ttaaggtacc gagggcctat ttcccatgat tccttcatat 2401 ttgcatatac gatacaaggc tgttagagag ataattagaa ttaatttgac tgtaaacaca 2461 aagatattag tacaaaatac gtgacgtaga aagtaataat ttcttgggta gtttgcagtt 2521 ttaaaattat gttttaaaat ggactatcat atgcttaccg taacttgaaa gtatttcgat 2581 ttcttggctt tatatatctt gtggaaagga cgaaacaccg gtgataagtg gaatgccatg 2641 gtttaagagc tatgctggaa acagcatagc aagtttaaat aaggctagtc cgttatcaac 2701 ttgaaaaagt ggcaccgagt cggtgctttt tttcctgcag cccgggggat ccactagttc 2761 tagagcggcc gccaccgcgg tggagctcca gcttttgttc cctttagtga gggttaattg 2821 cgcgcgaatt cgctagctag gtcttgaaag gagtgggaat tggctccggt gcccgtcagt 2881 gggcagagcg cacatcgccc acagtccccg agaagttggg gggaggggtc ggcaattgat 2941 ccggtgccta gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg tactggctcc 3001 gcctttttcc cgagggtggg ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc 3061 tttttcgcaa cgggtttgcc gccagaacac aggaccggtt ctagagcgct ctcgagagaa 3121 ccatgcagat ggagctcgaa atcaggccgc tgttcctcgt gccggacact aatggtttta 3181 tagatcactt ggcgtccttg gctagacttc tggaaagccg aaagtatata ttggtagtgc 3241 cgttgattgt aattaacgaa ttggatgggt tggcgaaagg acaagagact gatcacagag 3301 caggaggcta cgcgagggtc gtccaagaga aggcgcgaaa aagcatcgag ttcctggagc 3361 agcgatttga gagcagggac tcatgcctga gagccctcac gtcacggggg aacgagctgg 3421 agtccatcgc tttccgaagt gaagacatta cgggccaact tgggaataat gatgacctca 3481 tcttgtcctg ctgcctgcac tactgcaagg acaaggctaa ggacttcatg cctgcctcca 3541 aggaggagcc tatccgattg ttgagggaag tagtactttt gacggacgac cgcaacctcc 3601 gggtaaaggc gctgactcga aatgtcccag taagggatat accggcgttc cttacatggg 3661 ctcaagtagg gagtggaagt gagacaccgg gaacctcaga gagcgccacg ccagaaagcg 3721 acaagaagta cagcatcggc ctggccatcg gcaccaactc tgtgggctgg gccgtgatca 3781 ccgacgagta caaggtgccc agcaagaaat tcaaggtgct gggcaacacc gaccggcaca 3841 gcatcaagaa gaacctgatc ggcgccctgc tgttcgacag cggagaaaca gccgaggcca 3901 cccggctgaa gagaaccgcc agaagaagat acaccagacg gaagaaccgg atctgctatc 3961 tgcaagagat cttcagcaac gagatggcca aggtggacga cagcttcttc cacagactgg 4021 aagagtcctt cctggtggaa gaggataaga agcacgagcg gcaccccatc ttcggcaaca 4081 tcgtggacga ggtggcctac cacgagaagt accccaccat ctaccacctg agaaagaaac 4141 tggtggacag caccgacaag gccgacctgc ggctgatcta tctggccctg gcccacatga 4201 tcaagttccg gggccacttc ctgatcgagg gcgacctgaa ccccgacaac agcgacgtgg 4261 acaagctgtt catccagctg gtgcagacct acaaccagct gttcgaggaa aaccccatca 4321 acgccagcgg cgtggacgcc aaggccatcc tgtctgccag actgagcaag agcagacggc 4381 tggaaaatct gatcgcccag ctgcccggcg agaagaagaa tggcctgttc ggcaacctga 4441 ttgccctgag cctgggcctg acccccaact tcaagagcaa cttcgacctg gccgaggatg 4501 ccaaactgca gctgagcaag gacacctacg acgacgacct ggacaacctg ctggcccaga 4561 tcggcgacca gtacgccgac ctgtttctgg ccgccaagaa cctgtccgac gccatcctgc 4621 tgagcgacat cctgagagtg aacaccgaga tcaccaaggc ccccctgagc gcctctatga 4681 tcaagagata cgacgagcac caccaggacc tgaccctgct gaaagctctc gtgcggcagc 4741 agctgcctga gaagtacaaa gagattttct tcgaccagag caagaacggc tacgccggct 4801 acatcgatgg cggagccagc caggaagagt tctacaagtt catcaagccc atcctggaaa 4861 agatggacgg caccgaggaa ctgctcgtga agctgaacag agaggacctg ctgcggaagc 4921 agcggacctt cgacaacggc agcatccccc accagatcca cctgggagag ctgcacgcca 4981 ttctgcggcg gcaggaagat ttttacccat tcctgaagga caaccgggaa aagatcgaga 5041 agatcctgac cttccgcatc ccctactacg tgggccctct ggccagggga aacagcagat 5101 tcgcctggat gaccagaaag agcgaggaaa ccatcacccc ctggaacttc gaggaagtgg 5161 tggacaaggg cgccagcgcc cagagcttca tcgagcggat gaccaacttc gataagaacc 5221 tgcccaacga gaaggtgctg cccaagcaca gcctgctgta cgagtacttc accgtgtaca 5281 acgagctgac caaagtgaaa tacgtgaccg agggaatgag aaagcccgcc ttcctgagcg 5341 gcgagcagaa aaaagccatc gtggacctgc tgttcaagac caaccggaaa gtgaccgtga 5401 agcagctgaa agaggactac ttcaagaaaa tcgagtgctt cgactccgtg gaaatctccg 5461 gcgtggaaga tcggttcaac gcctccctgg gcacatacca cgatctgctg aaaattatca 5521 aggacaagga cttcctggac aatgaggaaa acgaggacat tctggaagat atcgtgctga 5581 ccctgacact gtttgaggac agagagatga tcgaggaacg gctgaaaacc tatgcccacc 5641 tgttcgacga caaagtgatg aagcagctga agcggcggag atacaccggc tggggcaggc 5701 tgagccggaa gctgatcaac ggcatccggg acaagcagtc cggcaagaca atcctggatt 5761 tcctgaagtc cgacggcttc gccaacagaa acttcatgca gctgatccac gacgacagcc 5821 tgacctttaa agaggacatc cagaaagccc aggtgtccgg ccagggcgat agcctgcacg 5881 agcacattgc caatctggcc ggcagccccg ccattaagaa gggcatcctg cagacagtga 5941 aggtggtgga cgagctcgtg aaagtgatgg gccggcacaa gcccgagaac atcgtgatcg 6001 aaatggccag agagaaccag accacccaga agggacagaa gaacagccgc gagagaatga 6061 agcggatcga agagggcatc aaagagctgg gcagccagat cctgaaagaa caccccgtgg 6121 aaaacaccca gctgcagaac gagaagctgt acctgtacta cctgcagaat gggcgggata 6181 tgtacgtgga ccaggaactg gacatcaacc ggctgtccga ctacgatgtg gacgctatcg 6241 tgcctcagag ctttctgaag gacgactcca tcgataacaa agtgctgact cggagcgaca 6301 agaaccgggg caagagcgac aacgtgccct ccgaagaggt cgtgaagaag atgaagaact 6361 actggcgcca gctgctgaat gccaagctga ttacccagag gaagttcgac aatctgacca 6421 aggccgagag aggcggcctg agcgaactgg ataaggccgg cttcatcaag agacagctgg 6481 tggaaacccg gcagatcaca aagcacgtgg cacagatcct ggactcccgg atgaacacta 6541 agtacgacga gaacgacaaa ctgatccggg aagtgaaagt gatcaccctg aagtccaagc 6601 tggtgtccga tttccggaag gatttccagt tttacaaagt gcgcgagatc aacaactacc 6661 accacgccca cgacgcctac ctgaacgccg tcgtgggaac cgccctgatc aaaaagtacc 6721 ctaagctgga aagcgagttc gtgtacggcg actacaaggt gtacgacgtg cggaagatga 6781 tcgccaagag cgagcaggaa atcggcaagg ctaccgccaa gtacttcttc tacagcaaca 6841 tcatgaactt tttcaagacc gagattaccc tggccaacgg cgagatccgg aagcggcctc 6901 tgatcgagac aaacggcgaa acaggcgaga tcgtgtggga taagggccgg gactttgcca 6961 ccgtgcggaa agtgctgtct atgccccaag tgaatatcgt gaaaaagacc gaggtgcaga 7021 caggcggctt cagcaaagag tctatcctgc ccaagaggaa cagcgacaag ctgatcgcca 7081 gaaagaagga ctgggaccct aagaagtacg gcggcttcga cagccccacc gtggcctatt 7141 ctgtgctggt ggtggccaaa gtggaaaagg gcaagtccaa gaaactgaag agtgtgaaag 7201 agctgctggg gatcaccatc atggaaagaa gcagcttcga gaagaatccc atcgactttc 7261 tggaagccaa gggctacaaa gaagtgaaaa aggacctgat catcaagctg cctaagtact 7321 ccctgttcga gctggaaaac ggccggaaga gaatgctggc ctctgccggc gaactgcaga 7381 agggaaacga actggccctg ccctccaaat atgtgaactt cctgtacctg gccagccact 7441 atgagaagct gaagggctcc cccgaggata atgagcagaa acagctgttt gtggaacagc 7501 acaaacacta cctggacgag atcatcgagc agatcagcga gttctccaag agagtgatcc 7561 tggccgacgc taatctggac aaggtgctga gcgcctacaa caagcacaga gacaagccta 7621 tcagagagca ggccgagaat atcatccacc tgtttaccct gaccaatctg ggagcccctg 7681 ccgccttcaa gtactttgac accaccatcg accggaagag gtacaccagc accaaagagg 7741 tgctggacgc caccctgatc caccagagca tcaccggcct gtacgagaca cggatcgacc 7801 tgtctcagct gggaggcgac gcctatccct atgacgtgcc cgattatgcc agcctgggca 7861 gcggctcccc caagaaaaaa cgcaaggtgg aagatcctaa gaaaaagcgg aaagtggacg 7921 tgttcgaagg gtccggcgca acaaacttct ctctgctgaa acaagccgga gatgtcgaag 7981 agaatcctgg accgaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc 8041 ccagggccgt acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg 8101 tcgatccgga ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg 8161 tcgggctcga catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga 8221 ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg cgcatggccg 8281 agttgagcgg ttcccggctg gccgcgcagc aacagatgga aggcctcctg gcgccgcacc 8341 ggcccaagga gcccgcgtgg ttcctggcca ccgtcggagt ctcgcccgac caccagggca 8401 agggtctggg cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc gccggggtgc 8461 ccgccttcct ggagacctcc gcgccccgca acctcccctt ctacgagcgg ctcggcttca 8521 ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc 8581 ccggtgcctg aacgcgttaa gtcgacaatc aacctctgga ttacaaaatt tgtgaaagat 8641 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc 8701 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct 8761 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca 8821 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt 8881 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg 8941 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga 9001 aatcatcgtc ctttccttgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt 9061 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc 9121 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt 9181 gggccgcctc cccgcgtcga ctttaagacc aatgacttac aaggcagctg tagatcttag 9241 ccacttttta aaagaaaagg ggggactgga agggctaatt cactcccaac gaagacaaga 9301 tctgcttttt gcttgtactg ggtctctctg gttagaccag atctgagcct gggagctctc 9361 tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt 9421 agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc 9481 agtgtggaaa atctctagca gggcccgttt aaacccgctg atcagcctcg actgtgcctt 9541 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg 9601 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt 9661 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca 9721 atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct 9781 ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg 9841 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt 9901 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc 9961 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg 10021 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg 10081 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct 10141 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg 10201 agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg 10261 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc 10321 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca 10381 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc 10441 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc 10501 cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct 10561 aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcagcacgt 10621 gttgacaatt aatcatcggc atagtatatc ggcatagtat aatacgacaa ggtgaggaac 10681 taaaccatgg ccaagttgac cagtgccgtt ccggtgctca ccgcgcgcga cgtcgccgga 10741 gcggtcgagt tctggaccga ccggctcggg ttctcccggg acttcgtgga ggacgacttc 10801 gccggtgtgg tccgggacga cgtgaccctg ttcatcagcg cggtccagga ccaggtggtg 10861 ccggacaaca ccctggcctg ggtgtgggtg cgcggcctgg acgagctgta cgccgagtgg 10921 tcggaggtcg tgtccacgaa cttccgggac gcctccgggc cggccatgac cgagatcggc 10981 gagcagccgt gggggcggga gttcgccctg cgcgacccgg ccggcaactg cgtgcacttc 11041 gtggccgagg agcaggactg acacgtgcta cgagatttcg attccaccgc cgccttctat 11101 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg 11161 gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac 11221 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt 11281 tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc 11341 tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca 11401 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg 11461 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg 11521 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc 11581 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta 11641 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag 11701 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg 11761 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg 11821 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg 11881 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga 11941 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc 12001 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt 12061 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact 12121 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg 12181 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt 12241 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt 12301 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct 12361 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg 12421 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt 12481 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt 12541 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc 12601 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg 12661 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc 12721 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg 12781 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca 12841 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga 12901 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct 12961 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg 13021 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca 13081 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata 13141 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct 13201 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact 13261 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa 13321 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc 13381 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga 13441 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga 13501 aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca 13561 gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg 13621 aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca 13681 attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca 13741 gatatacgcg tt //
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LentiRCas9Lambda2载体质粒应用举例
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