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&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    3721..3769
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(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
//
