LOCUS Cellecta-pRCCH-C 11573 bp ds-DNA circular SYN 30-SEP-2017 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 11573) AUTHORS Cellecta TITLE Direct Submission JOURNAL Exported Sep 30, 2017 from SnapGene 4.0.5 to Vector NTI(R) format http://www.snapgene.com COMMENT VNTDATE|704851200| COMMENT VNTDBDATE|818208000| COMMENT VNTNAME|Cellecta-pRCCH-CMV-Cas9-2A-Hygro.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 2082..2285 /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..7639 /codon_start=1 /label=HygroR misc_feature 7652..8240 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" LTR 8314..8547 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" polyA_signal 8619..8740 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" rep_origin 8780..8915 /label=SV40 ori /note="SV40 origin of replication" rep_origin complement(9350..9938) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(10109..10969) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" promoter complement(10970..11074) /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 aaaaagcctg aactcaccgc gacgtctgtc gagaagtttc tgatcgaaaa 6721 gttcgacagc gtctccgacc tgatgcagct ctcggagggc gaagaatctc gtgctttcag 6781 cttcgatgta ggagggcgtg gatatgtcct gcgggtaaat agctgcgccg atggtttcta 6841 caaagatcgt tatgtttatc ggcactttgc atcggccgcg ctcccgattc cggaagtgct 6901 tgacattggg gaattcagcg agagcctgac ctattgcatc tcccgccgtg cacagggtgt 6961 cacgttgcaa gacctgcctg aaaccgaact gcccgctgtt ctgcagccgg tcgcggaggc 7021 catggatgcg atcgctgcgg ccgatcttag ccagacgagc gggttcggcc cattcggacc 7081 gcaaggaatc ggtcaataca ctacatggcg tgatttcata tgcgcgattg ctgatcccca 7141 tgtgtatcac tggcaaactg tgatggacga caccgtcagt gcgtccgtcg cgcaggctct 7201 cgatgagctg atgctttggg ccgaggactg ccccgaagtc cggcacctcg tgcacgcgga 7261 tttcggctcc aacaatgtcc tgacggacaa tggccgcata acagcggtca ttgactggag 7321 cgaggcgatg ttcggggatt cccaatacga ggtcgccaac atcttcttct ggaggccgtg 7381 gttggcttgt atggagcagc agacgcgcta cttcgagcgg aggcatccgg agcttgcagg 7441 atcgccgcgg ctccgggcgt atatgctccg cattggtctt gaccaactct atcagagctt 7501 ggttgacggc aatttcgatg atgcagcttg ggcgcagggt cgatgcgacg caatcgtccg 7561 atccggagcc gggactgtcg ggcgtacaca aatcgcccgc agaagcgcgg ccgtctggac 7621 cgatggctgt gtagaagtct gataagtcga caatcaacct ctggattaca aaatttgtga 7681 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt 7741 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa 7801 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt 7861 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct 7921 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg 7981 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc 8041 ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg 8101 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct 8161 gctgccggct ctgcggcctc ttccgcgtct ccgccttcgc cctcagacga gtcggatctc 8221 tctttgggcc gcctccccgc ctggtacctt taagaccaat gacttacaag gcagctgtag 8281 atcttagcca ctttttaaaa gaaaaggggg gactggaagg gctaattcac tcccaacgaa 8341 gataagatct gctttttgct tgtactgggt ctctctggtt agaccagatc tgagcctggg 8401 agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg ccttgagtgc 8461 ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc ctcagaccct 8521 tttagtcagt gtggaaaatc tctagcagta gtagttcatg tcatcttatt attcagtatt 8581 tataacttgc aaagaaatga atatcagaga gtgagaggaa cttgtttatt gcagcttata 8641 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc 8701 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg ctctagctat 8761 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc 8821 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct 8881 attccagaag tagtgaggag gcttttttgg aggcctagac ttttgcagag accaaattcg 8941 taatcatgtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca 9001 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat 9061 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt 9121 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct 9181 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa 9241 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa 9301 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc 9361 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga 9421 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc 9481 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt 9541 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct 9601 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg 9661 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta 9721 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct 9781 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa 9841 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt 9901 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta 9961 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat 10021 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa 10081 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct 10141 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta 10201 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct 10261 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg 10321 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa 10381 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt 10441 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta 10501 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca 10561 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta 10621 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct 10681 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg 10741 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac 10801 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact 10861 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa 10921 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt 10981 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat 11041 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg 11101 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc 11161 cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg 11221 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt 11281 cagcgggtgt tggcgggtgt cggggctggc ttaactatgc ggcatcagag cagattgtac 11341 tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca 11401 tcaggcgcca ttcgccattc aggctgcgca actgttggga agggcgatcg gtgcgggcct 11461 cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa 11521 cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgccaag ctg //