LOCUS Cellecta-pRCCN-C 11402 bp ds-DNA circular SYN 17-JAN-2019 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 11402) AUTHORS Cellecta TITLE Direct Submission JOURNAL Exported Jan 17, 2019 from SnapGene 4.2.11 to Vector NTI(R) format http://www.snapgene.com COMMENT VNTDATE|728438400| COMMENT VNTDBDATE|866851200| COMMENT VNTNAME|Cellecta-pRCCN-CMV-Cas9-2A-Neo.dna| COMMENT VNTAUTHORNAME|Cellecta| 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." misc_feature 1818..1933 /label=cPPT /note="central polypurine tract and central termination sequence of HIV-1 (lacking the first T)" promoter 2005..2348 /label=CMV /note="human cytomegalovirus (CMV) immediate early promoter" CDS 2369..2389 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS CDS 2393..2413 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS CDS 2417..2437 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS CDS 2444..6544 /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 6545..6592 /codon_start=1 /product="bipartite nuclear localization signal from nucleoplasmin" /label=nucleoplasmin NLS CDS 6608..6661 /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 6668..7471 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" misc_feature 7481..8069 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" LTR 8143..8376 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" polyA_signal 8448..8569 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" rep_origin 8609..8744 /label=SV40 ori /note="SV40 origin of replication" rep_origin complement(9179..9767) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(9938..10798) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" promoter complement(10799..10903) /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 ttgcttggat cgagcaattt aaaagaaaag gggggattgg ggggtacagt gcaggggaaa 1861 gaatagtaga cataatagca acagacatac aaactaaaga attacaaaaa caaattacaa 1921 aaattcaaaa tttctgcgtt gttgtcggtg ctcgttctct gctcttcacg ctactgaatt 1981 catcaccggt tcttcgaatc tagtattatg cccagtacat gaccttatgg gactttccta 2041 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt 2101 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg 2161 acgccaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca 2221 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtggaaggtt tatataagca 2281 gagctcgttt agtgaaccgt cagatcgcct ggagacgcca tccacgctgt tttgacctcc 2341 atagaagatt ctagccgcca ccatggcccc aaaaaagaag agaaaggtag atccaaaaaa 2401 gaagagaaag gtagatccaa aaaagaagag aaaggtaggt ggagacaaga agtacagcat 2461 cggcctggac atcggcacca actctgtggg ctgggccgtg atcaccgacg agtacaaggt 2521 gcccagcaag aaattcaagg tgctgggcaa caccgaccgg cacagcatca agaagaacct 2581 gatcggagcc ctgctgttcg acagcggcga aacagccgag gccacccggc tgaagagaac 2641 cgccagaaga agatacacca gacggaagaa ccggatctgc tatctgcaag agatcttcag 2701 caacgagatg gccaaggtgg acgacagctt cttccacaga ctggaagagt ccttcctggt 2761 ggaagaggat aagaagcacg agcggcaccc catcttcggc aacatcgtgg acgaggtggc 2821 ctaccacgag aagtacccca ccatctacca cctgagaaag aaactggtgg acagcaccga 2881 caaggccgac ctgcggctga tctatctggc cctggcccac atgatcaagt tccggggcca 2941 cttcctgatc gagggcgacc tgaaccccga caacagcgac gtggacaagc tgttcatcca 3001 gctggtgcag acctacaacc agctgttcga ggaaaacccc atcaacgcca gcggcgtgga 3061 cgccaaggcc atcctgtctg ccagactgag caagagcaga cggctggaaa atctgatcgc 3121 ccagctgccc ggcgagaaga agaatggcct gttcggaaac ctgattgccc tgagcctggg 3181 cctgaccccc aacttcaaga gcaacttcga cctggccgag gatgccaaac tgcagctgag 3241 caaggacacc tacgacgacg acctggacaa cctgctggcc cagatcggcg accagtacgc 3301 cgacctgttt ctggccgcca agaacctgtc cgacgccatc ctgctgagcg acatcctgag 3361 agtgaacacc gagatcacca aggcccccct gagcgcctct atgatcaaga gatacgacga 3421 gcaccaccag gacctgaccc tgctgaaagc tctcgtgcgg cagcagctgc ctgagaagta 3481 caaagagatt ttcttcgacc agagcaagaa cggctacgcc ggctacattg acggcggagc 3541 cagccaggaa gagttctaca agttcatcaa gcccatcctg gaaaagatgg acggcaccga 3601 ggaactgctc gtgaagctga acagagagga cctgctgcgg aagcagcgga ccttcgacaa 3661 cggcagcatc ccccaccaga tccacctggg agagctgcac gccattctgc ggcggcagga 3721 agatttttac ccattcctga aggacaaccg ggaaaagatc gagaagatcc tgaccttccg 3781 catcccctac tacgtgggcc ctctggccag gggaaacagc agattcgcct ggatgaccag 3841 aaagagcgag gaaaccatca ccccctggaa cttcgaggaa gtggtggaca agggcgcttc 3901 cgcccagagc ttcatcgagc ggatgaccaa cttcgataag aacctgccca acgagaaggt 3961 gctgcccaag cacagcctgc tgtacgagta cttcaccgtg tataacgagc tgaccaaagt 4021 gaaatacgtg accgagggaa tgagaaagcc cgccttcctg agcggcgagc agaaaaaggc 4081 catcgtggac ctgctgttca agaccaaccg gaaagtgacc gtgaagcagc tgaaagagga 4141 ctacttcaag aaaatcgagt gcttcgactc cgtggaaatc tccggcgtgg aagatcggtt 4201 caacgcctcc ctgggcacat accacgatct gctgaaaatt atcaaggaca aggacttcct 4261 ggacaatgag gaaaacgagg acattctgga agatatcgtg ctgaccctga cactgtttga 4321 ggacagagag atgatcgagg aacggctgaa aacctatgcc cacctgttcg acgacaaagt 4381 gatgaagcag ctgaagcggc ggagatacac cggctggggc aggctgagcc ggaagctgat 4441 caacggcatc cgggacaagc agtccggcaa gacaatcctg gatttcctga agtccgacgg 4501 cttcgccaac agaaacttca tgcagctgat ccacgacgac agcctgacct ttaaagagga 4561 catccagaaa gcccaggtgt ccggccaggg cgatagcctg cacgagcaca ttgccaatct 4621 ggccggcagc cccgccatta agaagggcat cctgcagaca gtgaaggtgg tggacgagct 4681 cgtgaaagtg atgggccggc acaagcccga gaacatcgtg atcgaaatgg ccagagagaa 4741 ccagaccacc cagaagggac agaagaacag ccgcgagaga atgaagcgga tcgaagaggg 4801 catcaaagag ctgggcagcc agatcctgaa agaacacccc gtggaaaaca cccagctgca 4861 gaacgagaag ctgtacctgt actacctgca gaatgggcgg gatatgtacg tggaccagga 4921 actggacatc aaccggctgt ccgactacga tgtggaccat atcgtgcctc agagctttct 4981 gaaggacgac tccatcgaca acaaggtgct gaccagaagc gacaagaacc ggggcaagag 5041 cgacaacgtg ccctccgaag aggtcgtgaa gaagatgaag aactactggc ggcagctgct 5101 gaacgccaag ctgattaccc agagaaagtt cgacaatctg accaaggccg agagaggcgg 5161 cctgagcgaa ctggataagg ccggcttcat caagagacag ctggtggaaa cccggcagat 5221 cacaaagcac gtggcacaga tcctggactc ccggatgaac actaagtacg acgagaatga 5281 caagctgatc cgggaagtga aagtgatcac cctgaagtcc aagctggtgt ccgatttccg 5341 gaaggatttc cagttttaca aagtgcgcga gatcaacaac taccaccacg cccacgacgc 5401 ctacctgaac gccgtcgtgg gaaccgccct gatcaaaaag taccctaagc tggaaagcga 5461 gttcgtgtac ggcgactaca aggtgtacga cgtgcggaag atgatcgcca agagcgagca 5521 ggaaatcggc aaggctaccg ccaagtactt cttctacagc aacatcatga actttttcaa 5581 gaccgagatt accctggcca acggcgagat ccggaagcgg cctctgatcg agacaaacgg 5641 cgaaaccggg gagatcgtgt gggataaggg ccgggatttt gccaccgtgc ggaaagtgct 5701 gagcatgccc caagtgaata tcgtgaaaaa gaccgaggtg cagacaggcg gcttcagcaa 5761 agagtctatc ctgcccaaga ggaacagcga taagctgatc gccagaaaga aggactggga 5821 ccctaagaag tacggcggct tcgacagccc caccgtggcc tattctgtgc tggtggtggc 5881 caaagtggaa aagggcaagt ccaagaaact gaagagtgtg aaagagctgc tggggatcac 5941 catcatggaa agaagcagct tcgagaagaa tcccatcgac tttctggaag ccaagggcta 6001 caaagaagtg aaaaaggacc tgatcatcaa gctgcctaag tactccctgt tcgagctgga 6061 aaacggccgg aagagaatgc tggcctctgc cggcgaactg cagaagggaa acgaactggc 6121 cctgccctcc aaatatgtga acttcctgta cctggccagc cactatgaga agctgaaggg 6181 ctcccccgag gataatgagc agaaacagct gtttgtggaa cagcacaagc actacctgga 6241 cgagatcatc gagcagatca gcgagttctc caagagagtg atcctggccg acgctaatct 6301 ggacaaagtg ctgtccgcct acaacaagca ccgggataag cccatcagag agcaggccga 6361 gaatatcatc cacctgttta ccctgaccaa tctgggagcc cctgccgcct tcaagtactt 6421 tgacaccacc atcgaccgga agaggtacac cagcaccaaa gaggtgctgg acgccaccct 6481 gatccaccag agcatcaccg gcctgtacga gacacggatc gacctgtctc agctgggagg 6541 cgacaaaagg ccggcggcca cgaaaaaggc cggccaggca aaaaagaaaa aggcggccgc 6601 aggatgcgag ggcagaggaa gccttctaac atgcggtgac gtggaggaga atcccggccc 6661 ttccgggatg ggatcggcca ttgaacaaga tggattgcac gcaggttctc cggccgcttg 6721 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc 6781 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg 6841 tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt 6901 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg 6961 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat 7021 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca 7081 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca 7141 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa 7201 ggcgcgcatg cccgacggcg atgatctcgt cgtgacccat ggcgatgcct gcttgccgaa 7261 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc 7321 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga 7381 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc 7441 cttctatcgc cttcttgacg agttcttctg ataagtcgac aatcaacctc tggattacaa 7501 aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata 7561 cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc 7621 cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg 7681 tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac 7741 ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat 7801 cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt 7861 ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg ccacctggat 7921 tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc 7981 ccgcggcctg ctgccggctc tgcggcctct tccgcgtctc cgccttcgcc ctcagacgag 8041 tcggatctct ctttgggccg cctccccgcc tggtaccttt aagaccaatg acttacaagg 8101 cagctgtaga tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact 8161 cccaacgaag ataagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct 8221 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc 8281 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc 8341 tcagaccctt ttagtcagtg tggaaaatct ctagcagtag tagttcatgt catcttatta 8401 ttcagtattt ataacttgca aagaaatgaa tatcagagag tgagaggaac ttgtttattg 8461 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt 8521 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctggc 8581 tctagctatc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca 8641 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc 8701 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctagact tttgcagaga 8761 ccaaattcgt aatcatgtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc 8821 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct 8881 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc 8941 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt 9001 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag 9061 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca 9121 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt 9181 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc 9241 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct 9301 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg 9361 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca 9421 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact 9481 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta 9541 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta 9601 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct 9661 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt 9721 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga 9781 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca 9841 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat 9901 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg 9961 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt 10021 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag 10081 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc 10141 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag 10201 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca 10261 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa 10321 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga 10381 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata 10441 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca 10501 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg 10561 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg 10621 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg 10681 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag 10741 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac 10801 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca 10861 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag 10921 tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta 10981 tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc 11041 agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc 11101 agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc 11161 agattgtact gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa 11221 aataccgcat caggcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg 11281 tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa 11341 gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgccaagc 11401 tg //