LOCUS Exported 6784 bp ds-DNA circular SYN 29-JUN-2018 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6784) AUTHORS . TITLE Direct Submission JOURNAL Exported Jul 7, 2018 from SnapGene Viewer 4.1.9 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6784 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..141 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" promoter 181..392 /label=EF-1-alpha core promoter /note="core promoter for human elongation factor EF-1-alpha" regulatory 425..434 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" misc_feature 431..3686 /label=dSaCas9 with NLSs CDS 437..457 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 3638..3685 /codon_start=1 /product="bipartite nuclear localization signal from nucleoplasmin" /label=nucleoplasmin NLS /translation="KRPAATKKAGQAKKKK" CDS 3737..3922 /codon_start=1 /product="Kruppel-associated box (KRAB) transcriptional repression domain from the human zinc finger protein ZNF10 (Margolin et al., 1994)" /label=KRAB /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY QLTKPDVILRLEKGEEP" repeat_region 4047..4187 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" rep_origin 4262..4717 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" promoter 4999..5103 /gene="bla" /label=AmpR promoter CDS 5104..5964 /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" rep_origin 6135..6723 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" ORIGIN 1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc 61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca 121 actccatcac taggggttcc tgcggccgca cgcgttcgag tggctccggt gcccgtcagt 181 gggcagagcg cacatcgccc acagtccccg agaagttggg gggaggggtc ggcaattgaa 241 cgggtgccta gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg tactggctcc 301 gcctttttcc cgagggtggg ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc 361 tttttcgcaa cgggtttgcc gccagaacac aggtgtcgtg acgcgggaat tcggtaccac 421 cggtgccacc atggccccaa agaagaagcg gaaggtcggt atccacggag tcccagcagc 481 caagcggaac tacatcctgg gcctggccat cggcatcacc agcgtgggct acggcatcat 541 cgactacgag acacgggacg tgatcgatgc cggcgtgcgg ctgttcaaag aggccaacgt 601 ggaaaacaac gagggcaggc ggagcaagag aggcgccaga aggctgaagc ggcggaggcg 661 gcatagaatc cagagagtga agaagctgct gttcgactac aacctgctga ccgaccacag 721 cgagctgagc ggcatcaacc cctacgaggc cagagtgaag ggcctgagcc agaagctgag 781 cgaggaagag ttctctgccg ccctgctgca cctggccaag agaagaggcg tgcacaacgt 841 gaacgaggtg gaagaggaca ccggcaacga gctgtccacc aaagagcaga tcagccggaa 901 cagcaaggcc ctggaagaga aatacgtggc cgaactgcag ctggaacggc tgaagaaaga 961 cggcgaagtg cggggcagca tcaacagatt caagaccagc gactacgtga aagaagccaa 1021 acagctgctg aaggtgcaga aggcctacca ccagctggac cagagcttca tcgacaccta 1081 catcgacctg ctggaaaccc ggcggaccta ctatgaggga cctggcgagg gcagcccctt 1141 cggctggaag gacatcaaag aatggtacga gatgctgatg ggccactgca cctacttccc 1201 cgaggaactg cggagcgtga agtacgccta caacgccgac ctgtacaacg ccctgaacga 1261 cctgaacaat ctcgtgatca ccagggacga gaacgagaag ctggaatatt acgagaagtt 1321 ccagatcatc gagaacgtgt tcaagcagaa gaagaagccc accctgaagc agatcgccaa 1381 agaaatcctc gtgaacgaag aggatattaa gggctacaga gtgaccagca ccggcaagcc 1441 cgagttcacc aacctgaagg tgtaccacga catcaaggac attaccgccc ggaaagagat 1501 tattgagaac gccgagctgc tggatcagat tgccaagatc ctgaccatct accagagcag 1561 cgaggacatc caggaagaac tgaccaatct gaactccgag ctgacccagg aagagatcga 1621 gcagatctct aatctgaagg gctataccgg cacccacaac ctgagcctga aggccatcaa 1681 cctgatcctg gacgagctgt ggcacaccaa cgacaaccag atcgctatct tcaaccggct 1741 gaagctggtg cccaagaagg tggacctgtc ccagcagaaa gagatcccca ccaccctggt 1801 ggacgacttc atcctgagcc ccgtcgtgaa gagaagcttc atccagagca tcaaagtgat 1861 caacgccatc atcaagaagt acggcctgcc caacgacatc attatcgagc tggcccgcga 1921 gaagaactcc aaggacgccc agaaaatgat caacgagatg cagaagcgga accggcagac 1981 caacgagcgg atcgaggaaa tcatccggac caccggcaaa gagaacgcca agtacctgat 2041 cgagaagatc aagctgcacg acatgcagga aggcaagtgc ctgtacagcc tggaagccat 2101 ccctctggaa gatctgctga acaacccctt caactatgag gtggaccaca tcatccccag 2161 aagcgtgtcc ttcgacaaca gcttcaacaa caaggtgctc gtgaagcagg aagaagccag 2221 caagaagggc aaccggaccc cattccagta cctgagcagc agcgacagca agatcagcta 2281 cgaaaccttc aagaagcaca tcctgaatct ggccaagggc aagggcagaa tcagcaagac 2341 caagaaagag tatctgctgg aagaacggga catcaacagg ttctccgtgc agaaagactt 2401 catcaaccgg aacctggtgg ataccagata cgccaccaga ggcctgatga acctgctgcg 2461 gagctacttc agagtgaaca acctggacgt gaaagtgaag tccatcaatg gcggcttcac 2521 cagctttctg cggcggaagt ggaagtttaa gaaagagcgg aacaaggggt acaagcacca 2581 cgccgaggac gccctgatca ttgccaacgc cgatttcatc ttcaaagagt ggaagaaact 2641 ggacaaggcc aaaaaagtga tggaaaacca gatgttcgag gaaaagcagg ccgagagcat 2701 gcccgagatc gaaaccgagc aggagtacaa agagatcttc atcacccccc accagatcaa 2761 gcacattaag gacttcaagg actacaagta cagccaccgg gtggacaaga agcctaatag 2821 agagctgatt aacgacaccc tgtactccac ccggaaggac gacaagggca acaccctgat 2881 cgtgaacaat ctgaacggcc tgtacgacaa ggacaatgac aagctgaaaa agctgatcaa 2941 caagagcccc gaaaagctgc tgatgtacca ccacgacccc cagacctacc agaaactgaa 3001 gctgattatg gaacagtacg gcgacgagaa gaatcccctg tacaagtact acgaggaaac 3061 cgggaactac ctgaccaagt actccaaaaa ggacaacggc cccgtgatca agaagattaa 3121 gtattacggc aacaaactga acgcccatct ggacatcacc gacgactacc ccaacagcag 3181 aaacaaggtc gtgaagctgt ccctgaagcc ctacagattc gacgtgtacc tggacaatgg 3241 cgtgtacaag ttcgtgaccg tgaagaatct ggatgtgatc aaaaaagaaa actactacga 3301 agtgaatagc aagtgctatg aggaagctaa gaagctgaag aagatcagca accaggccga 3361 gtttatcgcc tccttctaca acaacgatct gatcaagatc aacggcgagc tgtatagagt 3421 gatcggcgtg aacaacgacc tgctgaaccg gatcgaagtg aacatgatcg acatcaccta 3481 ccgcgagtac ctggaaaaca tgaacgacaa gaggcccccc aggatcatta agacaatcgc 3541 ctccaagacc cagagcatta agaagtacag cacagacatt ctgggcaacc tgtatgaagt 3601 gaaatctaag aagcaccctc agatcatcaa aaagggcaaa aggccggcgg ccacgaaaaa 3661 ggccggccag gcaaaaaaga aaaagggatc cggcggtggc ggagggatgg atgctaagtc 3721 actaactgcc tggtcccgga cactggtgac cttcaaggat gtatttgtgg acttcaccag 3781 ggaggagtgg aagctgctgg acactgctca gcagatcgtg tacagaaatg tgatgctgga 3841 gaactataag aacctggttt ccttgggtta tcagcttact aagccagatg tgatcctccg 3901 gttggagaag ggagaagagc cctagtctag acatatggat atctgactag tctcgagttt 3961 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca 4021 tttttttcac tggaccgagc ggccgcagga acccctagtg atggagttgg ccactccctc 4081 tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt 4141 tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg 4201 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag 4261 tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc 4321 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc 4381 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt 4441 agtgctttac ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg 4501 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt 4561 ggactcttgt tccaaactgg aacaacactc aaccctatct cgggctattc ttttgattta 4621 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt 4681 aacgcgaatt ttaacaaaat attaacgttt acaattttat ggtgcactct cagtacaatc 4741 tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc 4801 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc 4861 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg 4921 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc 4981 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat 5041 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag 5101 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt 5161 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt 5221 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc 5281 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta 5341 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac 5401 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa 5461 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg 5521 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc 5581 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg 5641 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta 5701 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg 5761 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg 5821 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc 5881 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt 5941 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt 6001 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc 6061 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag 6121 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa 6181 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg 6241 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag 6301 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg 6361 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga 6421 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc 6481 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc 6541 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga 6601 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt 6661 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg 6721 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac 6781 atgt //