LOCUS       V034533                 6528 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V034533
VERSION     V034533
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"
     promoter        678..696
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /label="T7 promoter"
     CDS             756..1208
                     /label="Cav3(NM_007617)"
                     /note="Cav3(NM_007617)"
                     /gene="Cav3"
     CDS             1215..1280
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    1304..1892
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    1958..2006
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      2208..2636
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        2650..2979
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             3046..3840
                     /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    4016..4137
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(4186..4202)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    4210..4226
                     /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(4234..4264)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    4279..4300
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(4588..5176)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(5347..6207)
                     /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(6208..6312)
                     /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 gctctctggc taactagaga acccactgct
      661 tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg ctagcgttta
      721 aacttaagct tggtaccgag ctcgaattcg ccaccatgat gaccgaagag cacacggatc
      781 tggaagctcg gatcatcaag gacattcact gcaaggagat agacttggtg aaccgcgacc
      841 ccaagaacat caatgaggac attgtgaagg tagattttga agacgtgatt gcggagcccg
      901 agggcaccta cagcttcgac ggtgtatgga aggtgagctt caccacgttc accgtctcca
      961 agtactggtg ctaccgcctg ttgtctacac tgctgggtgt tccactggcc ctgctctggg
     1021 gattcctgtt cgcctgtatc tccttctgcc acatctgggc cgtggtgccc tgcattaaga
     1081 gctacctgat cgagatccag tgcatcagcc acatctactc actgtgtatc cgcaccttct
     1141 gcaacccgct cttcgctgcg ttgggccagg tctgcagcaa cattaaggtg gtgctgcgaa
     1201 gggaaggcgg atccgactac aaagaccatg acggtgatta taaagatcat gacatcgact
     1261 acaaggatga cgatgacaag tagtgagggc ccgatatctc gacaatcaac ctctggatta
     1321 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg
     1381 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc
     1441 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca
     1501 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac
     1561 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact
     1621 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc
     1681 cgtggtgttg tcggggaagc tgacgtcctt tccatggctg ctcgcctgtg ttgccacctg
     1741 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc
     1801 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac
     1861 gagtcggatc tccctttggg ccgcctcccc gcctggaaac gggggaggct aactgaaaca
     1921 cggaaggaga caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa
     1981 acgcacgggt gttgggtcgt ttgttcataa acgcggggtt cggtcccagg gctggcactc
     2041 tgtcgatacc ccaccgagac cccattgggg ccaatacgcc cgcgtttctt ccttttcccc
     2101 accccacccc ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg gggcggcagg
     2161 ccctgccata gcagatctgc gcagctgggg ctctaggggg tatccccacg cgccctgtag
     2221 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     2281 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     2341 tccccgtcaa gctctaaatc ggggcatccc tttagggttc cgatttagtg ctttacggca
     2401 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     2461 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     2521 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttggg
     2581 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt
     2641 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     2701 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     2761 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     2821 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     2881 ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag
     2941 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata
     3001 tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat
     3061 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     3121 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     3181 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg
     3241 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     3301 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     3361 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     3421 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     3481 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     3541 gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc
     3601 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     3661 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     3721 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     3781 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     3841 gcgggactct ggggttcgcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     3901 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     3961 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
     4021 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     4081 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     4141 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc
     4201 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     4261 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     4321 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     4381 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     4441 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     4501 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     4561 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     4621 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     4681 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     4741 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct caatgctcac gctgtaggta
     4801 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     4861 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     4921 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     4981 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg
     5041 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5101 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5161 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5221 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     5281 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     5341 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5401 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     5461 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     5521 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     5581 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     5641 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     5701 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     5761 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     5821 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     5881 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     5941 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6001 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6061 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6121 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6181 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6241 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6301 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat cgggagatct
     6361 cccgatcccc tatggtcgac tctcagtaca atctgctctg atgccgcata gttaagccag
     6421 tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa aatttaagct
     6481 acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagg
//
