LOCUS Cellecta-pRSGCCB 11304 bp ds-DNA circular SYN 11-AUG-2017 DEFINITION synthetic circular DNA. SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 11304) AUTHORS Sylvain Baron TITLE Direct Submission JOURNAL Exported Aug 11, 2017 from SnapGene 2.7.3 to Vector NTI(R) format http://www.snapgene.com COMMENT VNTDATE|758592000| COMMENT VNTDBDATE|813801600| COMMENT VNTNAME|Cellecta-pRSGCCB-U6-sg-HTS6C-CMV-Cas9-2A-Blast| COMMENT VNTAUTHORNAME|Sylvain Baron| FEATURES Location/Qualifiers promoter 6..232 /label=RSV promoter /note="Rous sarcoma virus enhancer/promoter" LTR 233..413 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 460..585 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1078..1311 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." promoter 1819..2059 /label=U6 /note="RNA polymerase III promoter for human U6 snRNA" misc_feature 2065..2094 /label=cloning site misc_feature 2122..2237 /label=cPPT /note="central polypurine tract and central termination sequence of HIV-1 (lacking the first T)" promoter 2309..2652 /label=CMV CDS 2673..2693 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS CDS 2697..2717 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS CDS 2721..2741 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS CDS 2748..6848 /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" CDS 6849..6896 /codon_start=1 /product="bipartite nuclear localization signal from nucleoplasmin" /label=nucleoplasmin NLS CDS 6912..6965 /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." CDS 6972..7370 /codon_start=1 /gene="Aspergillus terreus BSD" /product="blasticidin S deaminase" /label=BSD /note="confers resistance to blasticidin" misc_feature 7383..7971 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" LTR 8045..8278 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" polyA_signal 8350..8471 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" rep_origin 8511..8646 /label=SV40 ori /note="SV40 origin of replication" rep_origin complement(9081..9669) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(9840..10700) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" promoter complement(10701..10805) /gene="bla" /label=AmpR promoter ORIGIN 1 acgcgtgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga tgagttagca 61 acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag taaggtggta 121 cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac gaaccactga 181 attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacataa acgggtctct 241 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa 301 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc 361 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc 421 ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg caggactcgg 481 cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac gccaaaaatt 541 ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat taagcggggg 601 agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa 661 ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa tcctggcctg 721 ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc ccttcagaca 781 ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg tgtgcatcaa 841 aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga gcaaaacaaa 901 agtaagacca ccgcacagca agcggccact gatcttcaga cctggaggag gagatatgag 961 ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac cattaggagt 1021 agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag tgggaatagg 1081 agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag cgtcaatgac 1141 gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga acaatttgct 1201 gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca tcaagcagct 1261 ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc tggggatttg 1321 gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta gttggagtaa 1381 taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca gagaaattaa 1441 caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc aagaaaagaa 1501 tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt ttaacataac 1561 aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg taggtttaag 1621 aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt caccattatc 1681 gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa tagaagaaga 1741 aggtggagag agagacagag acagatccat tcgattagtg aacggatctc gacggtatcg 1801 atggccgccc ccttcaccga gggcctattt cccatgattc cttcatattt gcatatacga 1861 tacaaggctg ttagagagat aattggaatt aatttgactg taaacacaaa gatattagta 1921 caaaatacgt gacgtagaaa gtaataattt cttgggtagt ttgcagtttt aaaattatgt 1981 tttaaaatgg actatcatat gcttaccgta acttgaaagt atttcgattt cttggcttta 2041 tatatcttgt ggaaaggacg aaacaccgga gtcttctttt ttgaagacac ttcggactgt 2101 agaactctga acctcgagca atttaaaaga aaagggggga ttggggggta cagtgcaggg 2161 gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt 2221 acaaaaattc aaaatttctg cgttgttgtc ggtgctcgtt ctctgctctt cacgctactg 2281 aattcatcac cggttcttcg aatctagtat tatgcccagt acatgacctt atgggacttt 2341 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg 2401 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc 2461 attgacgcca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt 2521 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggaa ggtttatata 2581 agcagagctc gtttagtgaa ccgtcagatc gcctggagac gccatccacg ctgttttgac 2641 ctccatagaa gattctagcc gccaccatgg ccccaaaaaa gaagagaaag gtagatccaa 2701 aaaagaagag aaaggtagat ccaaaaaaga agagaaaggt aggtggagac aagaagtaca 2761 gcatcggcct ggacatcggc accaactctg tgggctgggc cgtgatcacc gacgagtaca 2821 aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc atcaagaaga 2881 acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc cggctgaaga 2941 gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg caagagatct 3001 tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa gagtccttcc 3061 tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc gtggacgagg 3121 tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg gtggacagca 3181 ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc aagttccggg 3241 gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac aagctgttca 3301 tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac gccagcggcg 3361 tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg gaaaatctga 3421 tcgcccagct gcccggcgag aagaagaatg gcctgttcgg aaacctgatt gccctgagcc 3481 tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc aaactgcagc 3541 tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc ggcgaccagt 3601 acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg agcgacatcc 3661 tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc aagagatacg 3721 acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag ctgcctgaga 3781 agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac attgacggcg 3841 gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag atggacggca 3901 ccgaggaact gctcgtgaag ctgaacagag aggacctgct gcggaagcag cggaccttcg 3961 acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt ctgcggcggc 4021 aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag atcctgacct 4081 tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc gcctggatga 4141 ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg gacaagggcg 4201 cttccgccca gagcttcatc gagcggatga ccaacttcga taagaacctg cccaacgaga 4261 aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtataac gagctgacca 4321 aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc gagcagaaaa 4381 aggccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag cagctgaaag 4441 aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc gtggaagatc 4501 ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag gacaaggact 4561 tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc ctgacactgt 4621 ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg ttcgacgaca 4681 aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg agccggaagc 4741 tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc ctgaagtccg 4801 acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg acctttaaag 4861 aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag cacattgcca 4921 atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag gtggtggacg 4981 agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa atggccagag 5041 agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag cggatcgaag 5101 agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa aacacccagc 5161 tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg tacgtggacc 5221 aggaactgga catcaaccgg ctgtccgact acgatgtgga ccatatcgtg cctcagagct 5281 ttctgaagga cgactccatc gacaacaagg tgctgaccag aagcgacaag aaccggggca 5341 agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac tggcggcagc 5401 tgctgaacgc caagctgatt acccagagaa agttcgacaa tctgaccaag gccgagagag 5461 gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg gaaacccggc 5521 agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag tacgacgaga 5581 atgacaagct gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg gtgtccgatt 5641 tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac cacgcccacg 5701 acgcctacct gaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct aagctggaaa 5761 gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc gccaagagcg 5821 agcaggaaat cggcaaggct accgccaagt acttcttcta cagcaacatc atgaactttt 5881 tcaagaccga gattaccctg gccaacggcg agatccggaa gcggcctctg atcgagacaa 5941 acggcgaaac cggggagatc gtgtgggata agggccggga ttttgccacc gtgcggaaag 6001 tgctgagcat gccccaagtg aatatcgtga aaaagaccga ggtgcagaca ggcggcttca 6061 gcaaagagtc tatcctgccc aagaggaaca gcgataagct gatcgccaga aagaaggact 6121 gggaccctaa gaagtacggc ggcttcgaca gccccaccgt ggcctattct gtgctggtgg 6181 tggccaaagt ggaaaagggc aagtccaaga aactgaagag tgtgaaagag ctgctgggga 6241 tcaccatcat ggaaagaagc agcttcgaga agaatcccat cgactttctg gaagccaagg 6301 gctacaaaga agtgaaaaag gacctgatca tcaagctgcc taagtactcc ctgttcgagc 6361 tggaaaacgg ccggaagaga atgctggcct ctgccggcga actgcagaag ggaaacgaac 6421 tggccctgcc ctccaaatat gtgaacttcc tgtacctggc cagccactat gagaagctga 6481 agggctcccc cgaggataat gagcagaaac agctgtttgt ggaacagcac aagcactacc 6541 tggacgagat catcgagcag atcagcgagt tctccaagag agtgatcctg gccgacgcta 6601 atctggacaa agtgctgtcc gcctacaaca agcaccggga taagcccatc agagagcagg 6661 ccgagaatat catccacctg tttaccctga ccaatctggg agcccctgcc gccttcaagt 6721 actttgacac caccatcgac cggaagaggt acaccagcac caaagaggtg ctggacgcca 6781 ccctgatcca ccagagcatc accggcctgt acgagacacg gatcgacctg tctcagctgg 6841 gaggcgacaa aaggccggcg gccacgaaaa aggccggcca ggcaaaaaag aaaaaggcgg 6901 ccgcaggatg cgagggcaga ggaagccttc taacatgcgg tgacgtggag gagaatcccg 6961 gccctcccgg gatggccaag cctttgtctc aagaagaatc caccctcatt gaaagagcaa 7021 cggctacaat caacagcatc cccatctctg aagattacag cgtcgccagc gcagctctct 7081 ctagcgacgg ccgcatcttc actggtgtca atgtatatca ttttactggg ggaccttgtg 7141 cagaactcgt ggtgctgggc actgctgctg ctgcggcagc tggcaacctg acttgtatcg 7201 tcgcgatcgg aaatgagaac aggggcatct tgagcccctg cggacggtgc cgacaggtgc 7261 ttctcgatct gcatcctggg atcaaagcca tagtgaagga cagtgatgga cagccgacgg 7321 cagttgggat tcgtgaattg ctgccctctg gttatgtgtg ggagggctaa tgataagtcg 7381 acaatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg 7441 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc 7501 gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt 7561 tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca 7621 ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc 7681 ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc 7741 tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc 7801 tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc 7861 tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc 7921 tccgccttcg ccctcagacg agtcggatct ctctttgggc cgcctccccg cctggtacct 7981 ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa agaaaagggg 8041 ggactggaag ggctaattca ctcccaacga agataagatc tgctttttgc ttgtactggg 8101 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg 8161 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt 8221 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt 8281 agtagttcat gtcatcttat tattcagtat ttataacttg caaagaaatg aatatcagag 8341 agtgagagga acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca 8401 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc 8461 aatgtatctt atcatgtctg gctctagcta tcccgcccct aactccgccc atcccgcccc 8521 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg 8581 cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg 8641 gaggcctaga cttttgcaga gaccaaattc gtaatcatgt catagctgtt tcctgtgtga 8701 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc 8761 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc 8821 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc 8881 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt 8941 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca 9001 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa 9061 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat 9121 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc 9181 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc 9241 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt 9301 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac 9361 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg 9421 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca 9481 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc 9541 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa 9601 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa 9661 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac 9721 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta 9781 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt 9841 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata 9901 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc 9961 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac 10021 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag 10081 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac 10141 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc 10201 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg 10261 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc 10321 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct 10381 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc 10441 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc 10501 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc 10561 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc 10621 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca 10681 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt 10741 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt 10801 ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca 10861 ttaacctata aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac 10921 ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat 10981 gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg 11041 cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatgcg gtgtgaaata 11101 ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc attcgccatt caggctgcgc 11161 aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg 11221 ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt 11281 aaaacgacgg ccagtgccaa gctg //