LOCUS Exported 9133 bp ds-DNA circular SYN 31-JAN-2018 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 9133) AUTHORS sa TITLE Direct Submission JOURNAL Exported Feb 6, 2018 from SnapGene 4.1.4 http://www.snapgene.com FEATURES Location/Qualifiers source 1..9133 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 132..335 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" promoter 377..395 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 409..432 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS /translation="MPKKKRKV" misc_feature 433..1119 /label=APOBEC misc_feature 700..702 /label=W90Y misc_feature 808..810 /label=R126E misc_feature 1168..4878 /label=LB CPF1 misc_feature 4891..5139 /label=UGI CDS 5152..5172 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 5179..5232 /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." /translation="EGRGSLLTCGDVEENPGP" CDS 5233..5481 /codon_start=1 /label=UGI /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML" CDS 5482..5502 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 5503..5559 /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 5560..5808 /codon_start=1 /label=UGI-2 /label=UGI /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML" CDS 5809..5829 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 5830..5883 /codon_start=1 /product="2A peptide from Thosea asigna virus capsid protein" /label=T2A-2 /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="EGRGSLLTCGDVEENPGP" CDS 5884..6132 /codon_start=1 /label=UGI-3 /label=UGI /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML" CDS 6145..6165 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS-2 /translation="PKKKRKV" polyA_signal 6194..6418 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" primer_bind complement(6489..6505) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" protein_bind 6513..6529 /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(6537..6567) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 6582..6603 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." rep_origin complement(6891..7479) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(7650..8510) /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" promoter complement(8511..8615) /gene="bla" /label=AmpR promoter enhancer 8885..131 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" ORIGIN 1 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg 61 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg 121 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact 181 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa 241 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta 301 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct 361 agagatccgc ggccgctaat acgactcact atagggagag ccgccaccat gcccaagaag 421 aagaggaaag tcatgagctc agagactggc ccagtggctg tggaccccac attgagacgg 481 cggatcgagc cccatgagtt tgaggtattc ttcgatccga gagagctccg caaggagacc 541 tgcctgcttt acgaaattaa ttgggggggc cggcactcca tttggcgaca tacatcacag 601 aacactaaca agcacgtcga agtcaacttc atcgagaagt tcacgacaga aagatatttc 661 tgtccgaaca caaggtgcag cattacctgg tttctcagct acagcccatg cggcgaatgt 721 agtagggcca tcactgaatt cctgtcaagg tatccccacg tcactctgtt tatttacatc 781 gcaaggctgt accaccacgc tgaccccgaa aatcgacaag gcctgcggga tttgatctct 841 tcaggtgtga ctatccaaat tatgactgag caggagtcag gatactgctg gagaaacttt 901 gtgaattata gcccgagtaa tgaagcccac tggcctaggt atccccatct gtgggtacga 961 ctgtacgttc ttgaactgta ctgcatcata ctgggcctgc ctccttgtct caacattctg 1021 agaaggaagc agccacagct gacattcttt accatcgctc ttcagtcttg tcattaccag 1081 cgactgcccc cacacattct ctgggccacc gggttgaaaa gcggcagcga gactcccggg 1141 acctcagagt ccgccacacc cgaaagtatg tcaaagctgg agaaattcac caactgttat 1201 agcctgtcta agaccctgcg cttcaaggca atcccagtgg gcaagacaca agagaacatt 1261 gacaacaaac ggctcctggt ggaggatgag aagagggctg aagattacaa gggcgttaag 1321 aagctgctgg ataggtacta tctgtcattc atcaacgatg tcctccacag tatcaagctg 1381 aagaatctga acaattacat ttctctgttc cggaagaaga cacggaccga gaaggagaac 1441 aaagagctgg agaatctgga gatcaacctg aggaaagaaa tagctaaggc tttcaaaggg 1501 aacgagggtt acaagtccct gttcaagaaa gacattatcg agactattct gcctgagttc 1561 ctggacgata aagatgagat cgccctcgtc aattccttca atgggtttac cacagccttt 1621 accggcttct tcgacaatag agagaatatg ttctctgaag aggccaaatc cactagcatc 1681 gcctttcgct gcataaacga gaacctgact aggtacatca gcaatatgga catctttgag 1741 aaagtcgatg ccatattcga caaacatgag gtgcaggaga ttaaggagaa gatcctgaac 1801 tcagattacg atgtcgaaga tttcttcgag ggagagttct tcaacttcgt gctcacacaa 1861 gagggcattg atgtgtacaa tgcaatcatt ggagggttcg tgacagagag tggcgagaag 1921 ataaagggcc tgaacgagta tatcaacctc tacaaccaga aaaccaagca gaaactgcct 1981 aagttcaagc cactgtacaa acaagtgctc tcagataggg aaagcctgag cttctacggt 2041 gaagggtata catcagatga agaagtgctc gaagtgttcc gcaacaccct caataagaac 2101 agtgaaatct tctcttcaat caagaagctg gagaaactgt tcaagaattt cgatgagtac 2161 tcctctgccg gaatctttgt gaagaatggc cctgcaatat ccactattag caaagacatc 2221 tttggcgagt ggaacgttat cagggataag tggaatgccg agtacgatga tattcatctc 2281 aagaagaaag ccgtggttac agagaaatac gaggatgata gacgcaagag ctttaagaag 2341 attggtagct tctctctcga acagctgcag gagtacgccg acgctgacct gtcagtcgtg 2401 gagaaactca aggagatcat aatccagaag gtggatgaaa tctacaaagt gtatggaagc 2461 tctgagaaac tcttcgatgc agactttgtt ctggagaaga gtctgaagaa gaacgacgca 2521 gtggttgcta tcatgaagga cctgctggat tctgttaagt ctttcgagaa ttacattaag 2581 gcattctttg gtgaagggaa ggagacaaat agggacgaga gcttctatgg cgactttgtt 2641 ctggcctacg acatcctcct caaggttgac cacatctatg acgctatacg gaattacgtt 2701 acccagaagc cctatagcaa agacaagttc aagctgtatt tccagaatcc acagtttatg 2761 ggtgggtggg ataaagacaa agaaacagat tacagggcca ctatcctgcg gtacggcagc 2821 aaatactatc tggctatcat ggataagaag tacgccaaat gcctccagaa gatcgacaag 2881 gacgacgtga acggtaacta cgagaagatc aattacaagc tcctgccagg acctaacaag 2941 atgctgccca aggtgttctt ctccaagaaa tggatggcct actataaccc aagcgaggac 3001 attcagaaga tatacaagaa tgggacattc aagaagggcg atatgttcaa cctcaacgac 3061 tgccacaagc tgattgattt cttcaaggat agcatttctc gctatcccaa gtggtctaat 3121 gcatacgatt tcaacttcag cgagactgag aagtacaaag acatcgctgg cttctaccgg 3181 gaggtggaag agcaaggcta taaggtgtca ttcgaatccg cttctaagaa ggaagtggat 3241 aagctcgtgg aagagggtaa gctgtacatg ttccagatat acaacaaaga cttcagcgat 3301 aagagccacg gcactccaaa cctccatact atgtatttca agctgctgtt tgacgagaac 3361 aaccacggac agattaggct gtcaggaggc gcagaactct tcatgcgcag agcttcactg 3421 aagaaggagg aactcgttgt ccacccagcc aatagcccta tagccaataa gaatccagac 3481 aatcctaaga aaaccactac tctgtcttac gatgtgtata aggataagag attctctgaa 3541 gatcagtacg aactgcacat acccattgcc attaacaagt gccctaagaa catcttcaag 3601 attaacacag aggttagagt gctcctgaaa cacgacgata acccttatgt tataggcatt 3661 gctcgcggag agagaaacct gctgtacatc gtcgtggtgg acggcaaagg caacatcgtg 3721 gaacagtaca gtctcaatga aatcattaac aatttcaacg gaatccgcat taagaccgac 3781 taccattctc tcctcgacaa gaaggagaaa gaaaggttcg aagcaagaca gaattggaca 3841 agtatagaga atatcaaaga actgaaggct gggtacatct ctcaggttgt gcacaagata 3901 tgtgagctgg tggagaagta cgacgctgtt atcgccctcg cggacctgaa tagcggcttc 3961 aagaactcca gggtgaaggt ggagaagcag gtgtatcaga agttcgagaa gatgctgatc 4021 gacaagctca actatatggt ggacaagaaa tccaatcctt gcgctactgg tggagccctg 4081 aagggctatc aaatcaccaa taagttcgaa tctttcaagt ctatgagcac ccagaatggc 4141 ttcatcttct acatacccgc atggctgaca tccaagattg atccctctac cggatttgtt 4201 aatctgctca agactaagta cacctctatt gctgactcaa agaagttcat atcatcattt 4261 gaccgcatca tgtacgtgcc agaagaggac ctgttcgagt ttgccctgga ttacaagaat 4321 ttctctcgga ctgacgccga ctacatcaag aagtggaagc tctactctta tggtaatcgg 4381 attcgcatat tccgcaatcc caagaagaat aacgtgttcg attgggagga agtttgcctc 4441 accagcgctt acaaggagct gttcaataag tatgggatta actaccagca gggcgacata 4501 agagccctgc tgtgcgaaca atctgataag gcattctatt cctctttcat ggcactgatg 4561 tcactgatgc tgcaaatgcg caattccatc accggaagaa cagacgtggc ctttctgatc 4621 tctcctgtca agaactcaga tggcatcttc tacgattccc gcaactatga agcacaggag 4681 aatgctatcc tgcctaagaa tgccgatgca aatggagcct ataacatcgc cagaaaggtc 4741 ctctgggcca taggacaatt caagaaagct gaagatgaga agctggacaa ggtgaagatc 4801 gccatttcaa acaaagagtg gctcgaatat gctcagacct cagtgaagca tggatcaccc 4861 aagaagaaac ggaaagtgtc tggtggttct actaatctgt cagatattat tgaaaaggag 4921 accggtaagc aactggttat ccaggaatcc atcctcatgc tcccagagga ggtggaagaa 4981 gtcattggga acaagccgga aagcgatata ctcgtgcaca ccgcctacga cgagagcacc 5041 gacgagaatg tcatgcttct gactagcgac gcccctgaat acaagccttg ggctctggtc 5101 atacaggata gcaacggtga gaacaagatt aagatgctct ctggtggttc tcccaagaag 5161 aagaggaaag tcgggcccga gggcagagga agtctgctaa catgcggtga cgtcgaggag 5221 aatcctggcc caactaatct gtcagatatt attgaaaagg agaccggtaa gcaactggtt 5281 atccaggaat ccatcctcat gctcccagag gaggtggaag aagtcattgg gaacaagccg 5341 gaaagcgata tactcgtgca caccgcctac gacgagagca ccgacgagaa tgtcatgctt 5401 ctgactagcg acgcccctga atacaagcct tgggctctgg tcatacagga tagcaacggt 5461 gagaacaaga ttaagatgct ccccaagaag aagaggaaag tcgctactaa cttcagcctg 5521 ctgaagcagg ctggagacgt ggaggagaac cctggaccta ccaacctgtc cgatatcatt 5581 gagaaagaga ccggcaaaca gctggtgatc caggagagca tcctgatgct gcccgaagag 5641 gtggaggaag tgatcggcaa caagcccgag tccgacatcc tggtgcacac agcctatgat 5701 gaatccaccg acgagaacgt gatgctgctg acctccgatg ctcccgagta taaaccctgg 5761 gcactggtga tccaggactc taatggagag aacaagatca agatgctgcc caagaagaag 5821 aggaaagtcg aaggacgggg ctccctcctg acctgtggcg atgtggaaga gaaccccggc 5881 cccacaaacc tcagtgacat tatcgagaag gaaacaggaa aacagctcgt cattcaagaa 5941 tctattctta tgttgcctga ggaagtcgaa gaggttattg gcaataaacc tgaatctgat 6001 attcttgtcc ataccgctta cgatgagtcc acagatgaaa atgttatgct gctcacatct 6061 gacgcaccag agtacaaacc atgggcgctc gttattcaag attccaacgg cgaaaacaaa 6121 atcaaaatgc tttctggtgg ttctcctaaa aaaaaaagaa aggtgtgagt ttaaacccgc 6181 tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg 6241 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt 6301 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc 6361 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct 6421 tctgaggcgg aaagaaccag ctggggctcg ataccgtcga cctctagcta gagcttggcg 6481 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac 6541 atacgagccg gaagcataaa gtgtaaagcc tagggtgcct aatgagtgag ctaactcaca 6601 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat 6661 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc 6721 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca 6781 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca 6841 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg 6901 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg 6961 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt 7021 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt 7081 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc 7141 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt 7201 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt 7261 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc 7321 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa 7381 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt 7441 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct 7501 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta 7561 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa 7621 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc 7681 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact 7741 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc 7801 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt 7861 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta 7921 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg 7981 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt 8041 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc 8101 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt 8161 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc 8221 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc 8281 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa 8341 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac 8401 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa 8461 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt 8521 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa 8581 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct 8641 gacgtcgacg gatcgggaga tcgatctccc gatcccctag ggtcgactct cagtacaatc 8701 tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct 8761 gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg 8821 aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg 8881 cgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat 8941 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg 9001 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata 9061 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta 9121 catcaagtgt atc //