LOCUS       V032665                 8784 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V032665
VERSION     V032665
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..3464
                     /label="Adam12(NM_007400)"
                     /note="Adam12(NM_007400)"
                     /gene="Adam12"
     CDS             3471..3536
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    3560..4148
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    4214..4262
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      4464..4892
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        4906..5235
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             5302..6096
                     /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    6272..6393
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(6442..6458)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6466..6482
                     /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(6490..6520)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6535..6556
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6844..7432)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7603..8463)
                     /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(8464..8568)
                     /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 ccaccatggc agagcgcccg gcgcggcgcg
      781 cgccccccgc ccgcgccctc ctgctggccc tggctggggc cctgctggcg ccccgtgcag
      841 cccgagggat gagtttgtgg gaccagagag gaacttacga agtggccaga gcctcccttc
      901 tgagcaagga ccctgggatc ccaggacaga gcatcccagc caaggatcat ccagacgtgc
      961 tgactgtgca actgcagctg gagagccgag acctgatcct cagcctggaa aggaatgagg
     1021 gactcattgc caatggcttc acggagaccc attatctgca agatggtact gatgtctctc
     1081 tcactcgaaa tcacacggat cattgttact accatggaca tgtgcaagga gatgctgcat
     1141 cagtggtcag cctcagtact tgctctggtc tccggggact tatcatgttt gaaaataaaa
     1201 cgtacagctt agagccaatg aaaaacacca ctgacagcta caaactcgtc ccagctgaga
     1261 gcatgacgaa catccaaggg ctgtgtgggt cacagcataa caagtccaac ctcaccatgg
     1321 aagatgtctc ccctggaacc tctcaaatgc gggcaagaag gcataagaga gagaccctta
     1381 agatgaccaa gtacgtagag ctggttattg tggcagacaa cagagagttt cagaggcaag
     1441 gaaaagacct ggagaaagtt aagcagcgat taatagagat cgccaatcac gttgacaagt
     1501 tttacagacc actgaacatc cggatcgtgc tggtaggagt ggaagtgtgg aatgacatcg
     1561 acaaatgctc tataagccag gacccattca ccagcctcca tgagtttcta gactggagaa
     1621 agataaagct tctacctcga aaatcccacg acaatgctca gcttatcagt ggggtttatt
     1681 tccaaggaac caccatcggc atggcaccca tcatgagcat gtgcactgca gaacagtctg
     1741 gaggagttgt catggaccat tcagacagcc cccttggtgc cgcagtgacc ttggcacatg
     1801 agctgggcca caacttcggg atgaaccatg acacactgga gaggggctgc agctgcagaa
     1861 tggccgcaga gaaaggaggc tgcatcatga acccgtccac ggggttccca ttccccatgg
     1921 tgttcagcag ctgcagcagg aaggacctgg aggctagcct ggagaagggc atggggatgt
     1981 gcctcttcaa cctaccagag gtcaagcagg cctttggggg ccggaagtgt ggaaatggct
     2041 atgtggaaga gggagaagag tgtgactgcg gagaaccgga ggaatgcacg aatcgctgct
     2101 gtaacgctac cacctgtact ctgaagccag atgctgtgtg cgcgcacggg cagtgctgtg
     2161 aagactgtca gctgaagcct ccaggaactg catgcagggg ctccagcaac tcctgtgacc
     2221 tcccagaatt ctgcacaggg actgcccctc actgtccagc caatgtgtac ctacatgatg
     2281 gccacccgtg tcagggcgtg gatggttact gctacaacgg catctgccag acccatgagc
     2341 agcagtgtgt cacgctctgg ggaccaggtg ctaaaccggc tcctggcatc tgctttgagc
     2401 gagtcaactc tgcaggagat ccttatggta actgtggcaa agactccaag agcgccttcg
     2461 ccaaatgtga gctgagagat gccaagtgtg ggaaaatcca gtgtcaaggt ggtgcaagcc
     2521 gacctgtcat tggtaccaat gctgtttcca tagaaacaaa tatcccacag caggaaggag
     2581 gtcggattct gtgccggggg acccatgtgt acttgggtga tgacatgcca gacccagggc
     2641 ttgtgcttgc aggaacaaag tgtgcagaag gaaaaatctg cctcaatcgt cgatgtcaga
     2701 atatcagtgt cttcggcgtt cacaagtgtg ccatgcagtg ccacggccga ggggtatgta
     2761 acaacaggaa gaattgccac tgtgaagccc actgggctcc acccttctgt gacaagtttg
     2821 gctttggagg aagcacagac agtggtccca tcaggcaagc agataaccag ggcttgactg
     2881 taggaatcct ggtgagcatc ctgtgtctgc ttgctgctgg atttgtggtg tatctcaaaa
     2941 ggaagacgtt gatgcggctg ctgttcacac ataaaaaaac caccatggaa aagctaaggt
     3001 gtgtgcaccc ttcccggaca cccagtggcc ctcaccttgg ccaggctcac cacacccccg
     3061 ggaaaggcct gctgatgaac cgggcaccac atttcaatac ccccaaggac aggcactcgc
     3121 tgaaatgcca gaacatggac atcagcaggc ccctcgacgc tcgagccgtc ccacagcttc
     3181 agtcacctca gcgagtgctc ctgcctctcc accagacccc acgtgcaccc agtggccctg
     3241 ccaggcccct gcccgccagt cctgcagtca ggcaggccca gggcattcga aaacccagtc
     3301 ctcctcagaa gcctctgcct gctgatccac tgagcaggac ttctcggctc actagtgcct
     3361 tggtgaggac cccagggcag caggaacctg ggcaccgccc agcccccatc agacctgccc
     3421 ctaagcatca agtacccaga ccttcccaca atgcctatat caagggatcc gactacaaag
     3481 accatgacgg tgattataaa gatcatgaca tcgactacaa ggatgacgat gacaagtagt
     3541 gagggcccga tatctcgaca atcaacctct ggattacaaa atttgtgaaa gattgactgg
     3601 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
     3661 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct
     3721 gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt
     3781 tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac
     3841 tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg
     3901 ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaagctgac
     3961 gtcctttcca tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg
     4021 ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct
     4081 gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc
     4141 ctccccgcct ggaaacgggg gaggctaact gaaacacgga aggagacaat accggaagga
     4201 acccgcgcta tgacggcaat aaaaagacag aataaaacgc acgggtgttg ggtcgtttgt
     4261 tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca
     4321 ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga
     4381 aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcag atctgcgcag
     4441 ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     4501 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     4561 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     4621 catcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     4681 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     4741 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     4801 ctcggtctat tcttttgatt tataagggat tttggggatt tcggcctatt ggttaaaaaa
     4861 tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg
     4921 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     4981 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     5041 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     5101 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     5161 gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     5221 ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag
     5281 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc
     5341 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     5401 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     5461 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg
     5521 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     5581 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     5641 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     5701 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     5761 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     5821 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     5881 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     5941 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     6001 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     6061 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgcgaaat
     6121 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     6181 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     6241 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     6301 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     6361 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     6421 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     6481 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     6541 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     6601 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     6661 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     6721 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     6781 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     6841 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     6901 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     6961 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     7021 aagcgtggcg ctttctcaat gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     7081 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     7141 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     7201 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     7261 gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt
     7321 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     7381 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     7441 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     7501 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     7561 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     7621 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     7681 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     7741 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     7801 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     7861 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     7921 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     7981 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     8041 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     8101 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     8161 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     8221 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     8281 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     8341 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     8401 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     8461 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     8521 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     8581 aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg gtcgactctc
     8641 agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg
     8701 gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac
     8761 aattgcatga agaatctgct tagg
//
