LOCUS V018590 8291 bp DNA circular SYN 01-JAN-1980 DEFINITION synthetic circular DNA. ACCESSION V018590 VERSION V018590 KEYWORDS . SOURCE . ORGANISM . . FEATURES Location/Qualifiers enhancer 50..429 /note="human cytomegalovirus immediate early enhancer" /label="CMV enhancer" promoter 430..633 /note="human cytomegalovirus (CMV) immediate early promoter" /label="CMV promoter" CDS 665..2962 /label="BRAF(NM_004333)" /note="BRAF(NM_004333)" /gene="BRAF" CDS 2978..3043 /codon_start=1 /product="three tandem FLAG® epitope tags, followed by an enterokinase cleavage site" /transl_table=1 /translation="DYKDHDGDYKDHDIDYKDDDDK" /label="3xFLAG" misc_feature 3067..3655 /note="woodchuck hepatitis virus posttranscriptional regulatory element" /label="WPRE" polyA_signal 3721..3769 /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" /label="HSV TK poly(A) signal" rep_origin 3971..4399 /direction=RIGHT /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" /label="f1 ori" promoter 4413..4742 /note="SV40 enhancer and early promoter" /label="SV40 promoter" CDS 4809..5603 /codon_start=1 /gene="aph(3')-II (or nptII)" /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin®)" /product="aminoglycoside phosphotransferase from Tn5" /transl_table=1 /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*" /label="NeoR/KanR" polyA_signal 5779..5900 /note="SV40 polyadenylation signal" /label="SV40 poly(A) signal" primer_bind complement(5949..5965) /note="common sequencing primer, one of multiple similar variants" /label="M13 rev" protein_bind 5973..5989 /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-β-D-thiogalactopyranoside (IPTG)." /label="lac operator" promoter complement(5997..6027) /note="promoter for the E. coli lac operon" /label="lac promoter" protein_bind 6042..6063 /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." /label="CAP binding site" rep_origin complement(6351..6939) /direction=LEFT /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" /label="ori" CDS complement(7110..7970) /codon_start=1 /gene="bla" /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /product="β-lactamase" /transl_table=1 /translation="MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGY IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEY SPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDR WEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRS ALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGA SLIKHW*" /label="AmpR" promoter complement(7971..8075) /gene="bla" /label="AmpR promoter" ORIGIN 1 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta 61 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag 121 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc 181 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga 241 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat 301 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc 361 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct 421 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca 481 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat 541 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg 601 cgtgtacggt gggaggtcta tataagcaga gctctggcta gcgtttaaac ttaagcttgc 661 caccatggcg gcgctgagcg gtggcggtgg tggcggcgcg gagccgggcc aggctctgtt 721 caacggggac atggagcccg aggccggcgc cggcgccggc gccgcggcct cttcggctgc 781 ggaccctgcc attccggagg aggtgtggaa tatcaaacaa atgattaagt tgacacagga 841 acatatagag gccctattgg acaaatttgg tggggagcat aatccaccat caatatatct 901 ggaggcctat gaagaataca ccagcaagct agatgcactc caacaaagag aacaacagtt 961 attggaatct ctggggaacg gaactgattt ttctgtttct agctctgcat caatggatac 1021 cgttacatct tcttcctctt ctagcctttc agtgctacct tcatctcttt cagtttttca 1081 aaatcccaca gatgtggcac ggagcaaccc caagtcacca caaaaaccta tcgttagagt 1141 cttcctgccc aacaaacaga ggacagtggt acctgcaagg tgtggagtta cagtccgaga 1201 cagtctaaag aaagcactga tgatgagagg tctaatccca gagtgctgtg ctgtttacag 1261 aattcaggat ggagagaaga aaccaattgg ttgggacact gatatttcct ggcttactgg 1321 agaagaattg catgtggaag tgttggagaa tgttccactt acaacacaca actttgtacg 1381 aaaaacgttt ttcaccttag cattttgtga cttttgtcga aagctgcttt tccagggttt 1441 ccgctgtcaa acatgtggtt ataaatttca ccagcgttgt agtacagaag ttccactgat 1501 gtgtgttaat tatgaccaac ttgatttgct gtttgtctcc aagttctttg aacaccaccc 1561 aataccacag gaagaggcgt ccttagcaga gactgcccta acatctggat catccccttc 1621 cgcacccgcc tcggactcta ttgggcccca aattctcacc agtccgtctc cttcaaaatc 1681 cattccaatt ccacagccct tccgaccagc agatgaagat catcgaaatc aatttgggca 1741 acgagaccga tcctcatcag ctcccaatgt gcatataaac acaatagaac ctgtcaatat 1801 tgatgacttg attagagacc aaggatttcg tggtgatgga ggatcaacca caggtttgtc 1861 tgctaccccc cctgcctcat tacctggctc actaactaac gtgaaagcct tacagaaatc 1921 tccaggacct cagcgagaaa ggaagtcatc ttcatcctca gaagacagga atcgaatgaa 1981 aacacttggt agacgggact cgagtgatga ttgggagatt cctgatgggc agattacagt 2041 gggacaaaga attggatctg gatcatttgg aacagtctac aagggaaagt ggcatggtga 2101 tgtggcagtg aaaatgttga atgtgacagc acctacacct cagcagttac aagccttcaa 2161 aaatgaagta ggagtactca ggaaaacacg acatgtgaat atcctactct tcatgggcta 2221 ttccacaaag ccacaactgg ctattgttac ccagtggtgt gagggctcca gcttgtatca 2281 ccatctccat atcattgaga ccaaatttga gatgatcaaa cttatagata ttgcacgaca 2341 gactgcacag ggcatggatt acttacacgc caagtcaatc atccacagag acctcaagag 2401 taataatata tttcttcatg aagacctcac agtaaaaata ggtgattttg gtctagctac 2461 agagaaatct cgatggagtg ggtcccatca gtttgaacag ttgtctggat ccattttgtg 2521 gatggcacca gaagtcatca gaatgcaaga taaaaatcca tacagctttc agtcagatgt 2581 atatgcattt ggaattgttc tgtatgaatt gatgactgga cagttacctt attcaaacat 2641 caacaacagg gaccagataa tttttatggt gggacgagga tacctgtctc cagatctcag 2701 taaggtacgg agtaactgtc caaaagccat gaagagatta atggcagagt gcctcaaaaa 2761 gaaaagagat gagagaccac tctttcccca aattctcgcc tctattgagc tgctggcccg 2821 ctcattgcca aaaattcacc gcagtgcatc agaaccctcc ttgaatcggg ctggtttcca 2881 aacagaggat tttagtctat atgcttgtgc ttctccaaaa acacccatcc aggcaggggg 2941 atatggtgcg tttcctgtcc acacgcggcc gctcgaggac tacaaagacc atgacggtga 3001 ttataaagat catgacatcg actacaagga tgacgatgac aagtagtgag ggcccgatat 3061 ctcgacaatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat 3121 gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct 3181 tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag 3241 gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc 3301 cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc 3361 ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct 3421 cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga agctgacgtc ctttccatgg 3481 ctgctcgcct gtgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg 3541 gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg 3601 cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcctgga 3661 aacgggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc cgcgctatga 3721 cggcaataaa aagacagaat aaaacgcacg ggtgttgggt cgtttgttca taaacgcggg 3781 gttcggtccc agggctggca ctctgtcgat accccaccga gaccccattg gggccaatac 3841 gcccgcgttt cttccttttc cccaccccac cccccaagtt cgggtgaagg cccagggctc 3901 gcagccaacg tcggggcggc aggccctgcc atagcagatc tgcgcagctg gggctctagg 3961 gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc 4021 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc 4081 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcggggcat ccctttaggg 4141 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca 4201 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc 4261 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct 4321 tttgatttat aagggatttt ggggatttcg gcctattggt taaaaaatga gctgatttaa 4381 caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc 4441 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt 4501 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt 4561 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg 4621 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct 4681 ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca 4741 aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca ggatgaggat 4801 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga 4861 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc 4921 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga 4981 atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg 5041 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc 5101 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg 5161 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga 5221 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc 5281 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca 5341 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg 5401 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct 5461 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg 5521 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc 5581 gccttcttga cgagttcttc tgagcgggac tctggggttc gcgaaatgac cgaccaagcg 5641 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc 5701 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg 5761 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat 5821 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc 5881 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg 5941 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac 6001 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca 6061 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat 6121 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc 6181 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca 6241 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca 6301 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg 6361 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg 6421 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt 6481 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt 6541 tctcaatgct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc 6601 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt 6661 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt 6721 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc 6781 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa 6841 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt 6901 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct 6961 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta 7021 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa 7081 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc 7141 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact 7201 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc 7261 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt 7321 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta 7381 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg 7441 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt 7501 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc 7561 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt 7621 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc 7681 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc 7741 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa 7801 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac 7861 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa 7921 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt 7981 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa 8041 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct 8101 gacgtcgacg gatcgggaga tctcccgatc ccctatggtc gactctcagt acaatctgct 8161 ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt 8221 agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga 8281 atctgcttag g //