LOCUS       V023458                 9024 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V023458
VERSION     V023458
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..3704
                     /label="Nbr1(NM_001024765)"
                     /note="Nbr1(NM_001024765)"
                     /gene="Nbr1"
     CDS             3711..3776
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    3800..4388
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    4454..4502
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      4704..5132
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        5146..5475
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             5542..6336
                     /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    6512..6633
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(6682..6698)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6706..6722
                     /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(6730..6760)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6775..6796
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7084..7672)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7843..8703)
                     /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(8704..8808)
                     /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 ctcggatccg ccaccatgga accacaggtt actctaaatg
      781 tgacttttaa aaatgaaact caaagctttc tggtttcgga tccagaaaat acaacttggg
      841 ctgatgttga agctatggta aaagtttcat ttgatctgaa tactatccaa ataaaatacc
      901 tggatgagga aaatgaggag atatccatca atagtcaagg agaatatgaa gaagcactta
      961 agatggctaa cattaagcaa ggaaatcaac tacagatgca agtccacgaa gggtgccatg
     1021 ttgtagatga agtgctcccc caaactgtag tagaaaaaca agcaactgcc aggacaggga
     1081 agaagccact cgcacattac tcttcgctgg taagagtctt gggatcagat atgaagacca
     1141 cagaggagcc tacagcagag gcacgttcac ctgttccttg tgacaccgac aagcctcaag
     1201 acaagccccc agactggttt acaagctacc tggagatgtt cagagaacaa gtggtgaagg
     1261 aaacggtgga gaagcttgaa cagaggttgc aggagaagct tgtcctccag aagccaccct
     1321 tcagttcctc accctcagaa gtctcaatgc ctatttcaga ggaggaaacc ctgtttttgc
     1381 cagagaacca gttcagctgg catattgctt gtagccactg ccaaaagagg atcgtcggtg
     1441 tgcgctacca gtgtagcctc tgcccatcct acaacatttg tgaagattgt gaagctggac
     1501 cttactccca tgacactaac cacattttgc taaaatttcg gagacctgtt gtgatttcct
     1561 ctgagccatt tttctactca aagtatccca ctcctcgtct gcctgctgct ttggaacaag
     1621 tcaggctcca aaagcaggtg gataagaatt ttgtgaaagc agaaaagcaa aggttgcggg
     1681 ctgagaagaa acagcggaag gcagaggtca aagagcttaa aaagcagctg aaactccacc
     1741 ggaagattca tttgtggaat tcaatccatg gactccagag tcccaagtcg cctttgggtc
     1801 gacctgagag cttgctgcag tctaacaccc tcatgcttcc tttgcagccc tgtgccccag
     1861 ttatgccaac actcagtgca gcattcgtgg atgagaattt gcctgatggg actcatcttc
     1921 agccaggaac caagtttatc aaacactgga ggatgaaaaa cacagggaat gtgaagtgga
     1981 gcgcagacac aaagctcaag ttcatgtggg gaaacttgac tctggcttct acagagaaga
     2041 aggatgtttt ggttccctgc ttgaaggctg gacacatcgg ggtggtgtct gtggagttca
     2101 tcgccccaac cttggagggg acatacactt tgcattggcg tctctctcac aaaggccagc
     2161 agtttgggcc tcgggtctgg tgcagtatca tagtggatcc ttttccttcc tctgagagcc
     2221 ctgttaacct tgaaagggat gggattagct caagcaaagc tgatgacttc acctgtgagc
     2281 aagaggaagc ttttcttcta gctgaagaag agattccatt gggtgaggtg acaaagcaga
     2341 cagaagggac aggatccagt gccccacaga agacacaaca tgctgccagt gagagggagc
     2401 tctacattcc atccgtggac cttctgactg cccaggacct gctgtccttt gaactgctgg
     2461 atataaacat tgttcaagaa ttggagagag tgccacacaa cacccctgtg gatgtgactc
     2521 cccgcatgtc tcctctgccg catgacagcc ctttaataga gaagccaggc ttggggcgga
     2581 tacaggaaga gagtgaaggg gcgggattga aagcatctcc tgattccact gtattaacaa
     2641 agagaaaggc cgagacccct gcttcagtcg aagaaacgga ggaagacctg agtgggaccc
     2701 aatttgtgtg tgagactgta atccgatccc taaccttgga tgctgctcca gaccacaacc
     2761 ctccttgcag gcagaggtcc ccacagaggg aattacagct gtattccaca gaagaacagc
     2821 agcctcttat gctgcccgga ttctgcagaa aggattcttc cttgaaattt gccttgcctg
     2881 aagaaggacc acatggagat gagagggagg agattgtcca cattgctgag gaagaagcca
     2941 ttgaggaaga ggatgtccaa gatgaagaag ttcagagtca gtcttctgct tcctctgaag
     3001 attatatcat catcctgcct gagtgctttg ataccagccg ccccctgggg gactccatgt
     3061 acagctctgc gctctcccag ccaggcctgg agcgaggggc tgaaggtgaa cctgggatcg
     3121 agtctgggca ggaaccagct gaggctagag agagactccc ggaaagggag agccagccaa
     3181 aggagcagag catcagtgac atcctcacga cctcacagca tctggacaca gtgcctctag
     3241 tcccagaagt ggcaggactt ccagcagcac tgtccaggag cgccccttgt ggacagtatg
     3301 aggcaccacg agtggattca ccagttacca tccaagaagt tctcccagtc cccgatcacg
     3361 tcagaggaga gcccagaggc tcaacaggac tcgcaaacag ccgacagaag agctgtgacc
     3421 actcaaggca ccacaatggg agcagcattg ctggaggact ggtgaaaggg gctttgtctg
     3481 ttgctgcctc tgcgtacaag gccctgtttt ctgggccacc agtcactgca cagcccgtca
     3541 tttctgaaga tcagaccgca gccctgatgg cccatctctt tgaaatggga ttctgcgaca
     3601 ggcagctgaa cctaagactg ctgagaaaac acaatcacaa tatcctgcag gttgtgacag
     3661 agctcctcca agtgaacaac aacgactggt acagccaccg ctatctcgag gactacaaag
     3721 accatgacgg tgattataaa gatcatgaca tcgactacaa ggatgacgat gacaagtagt
     3781 gagggcccga tatctcgaca atcaacctct ggattacaaa atttgtgaaa gattgactgg
     3841 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
     3901 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct
     3961 gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt
     4021 tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac
     4081 tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg
     4141 ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaagctgac
     4201 gtcctttcca tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg
     4261 ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct
     4321 gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc
     4381 ctccccgcct ggaaacgggg gaggctaact gaaacacgga aggagacaat accggaagga
     4441 acccgcgcta tgacggcaat aaaaagacag aataaaacgc acgggtgttg ggtcgtttgt
     4501 tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca
     4561 ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga
     4621 aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcag atctgcgcag
     4681 ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     4741 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     4801 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     4861 catcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     4921 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     4981 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     5041 ctcggtctat tcttttgatt tataagggat tttggggatt tcggcctatt ggttaaaaaa
     5101 tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg
     5161 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     5221 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     5281 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     5341 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     5401 gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     5461 ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag
     5521 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc
     5581 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     5641 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     5701 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg
     5761 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     5821 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     5881 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     5941 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     6001 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     6061 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     6121 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     6181 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     6241 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     6301 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgcgaaat
     6361 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     6421 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     6481 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     6541 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     6601 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     6661 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     6721 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     6781 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     6841 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     6901 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     6961 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     7021 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     7081 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     7141 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     7201 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     7261 aagcgtggcg ctttctcaat gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     7321 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     7381 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     7441 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     7501 gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt
     7561 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     7621 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     7681 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     7741 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     7801 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     7861 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     7921 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     7981 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     8041 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     8101 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     8161 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     8221 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     8281 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     8341 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     8401 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     8461 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     8521 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     8581 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     8641 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     8701 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     8761 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     8821 aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg gtcgactctc
     8881 agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg
     8941 gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac
     9001 aattgcatga agaatctgct tagg
//
