LOCUS       V030483                 7835 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V030483
VERSION     V030483
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     enhancer        202..581
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        582..785
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     promoter        830..848
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /label="T7 promoter"
     regulatory      902..911
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             911..976
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             977..3328
                     /label="PFKP(NM_002627)"
                     /note="PFKP(NM_002627)"
                     /gene="PFKP"
     CDS             3338..3355
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /transl_table=1
                     /translation="HHHHHH"
                     /label="6xHis"
     polyA_signal    3402..3626
                     /note="bovine growth hormone polyadenylation signal"
                     /label="bGH poly(A) signal"
     rep_origin      3672..4100
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        4114..4443
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             4510..5304
                     /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    5478..5599
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(5648..5664)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    5672..5688
                     /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(5696..5726)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    5741..5762
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6050..6635)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(6806..7666)
                     /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(7667..7771)
                     /gene="bla"
                     /label="AmpR promoter"
ORIGIN
        1 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
       61 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
      121 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
      181 tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta
      241 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
      301 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
      361 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
      421 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
      481 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
      541 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
      601 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
      661 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
      721 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
      781 gagctctctg gctaactaga gaacccactg cttactggct tatcgaaatt aatacgactc
      841 actataggga gacccaagct ggctagcgtt taaacttaag cttggtaccg agctcggatc
      901 cgccaccatg gactacaaag accatgacgg tgattataaa gatcatgaca tcgactacaa
      961 ggatgacgat gacaagatgg acgcggacga ctcccgggcc cccaagggct ccttgcggaa
     1021 gttcctggag cacctctccg gggccggcaa ggccatcggc gtgctgacca gcggcgggga
     1081 tgctcaaggt atgaacgctg ccgtccgtgc cgtggtgcgc atgggtatct acgtgggggc
     1141 caaggtgtac ttcatctacg agggctacca gggcatggtg gacggaggct caaacatcgc
     1201 agaggccgac tgggagagtg tctccagcat cctgcaagtg ggcgggacga tcattggcag
     1261 tgcgcggtgc caggccttcc gcacgcggga aggccgcctg aaggctgctt gcaacctgct
     1321 gcagcgcggc atcaccaacc tgtgtgtgat cggcggggac gggagcctca ccggggccaa
     1381 cctcttccgg aaggagtgga gtgggctgct ggaggagctg gccaggaacg gccagatcga
     1441 taaggaggcc gtgcagaagt acgcctacct caacgtggtg ggcatggtgg gctccatcga
     1501 caatgatttc tgcggcaccg acatgaccat cggcacggac tccgccctgc acaggatcat
     1561 cgaggtcgtc gacgccatca tgaccacggc ccagagccac cagaggacct tcgttctgga
     1621 ggtgatggga cgacactgtg ggtacctggc cctggtgagt gccttggcct gcggtgcgga
     1681 ctgggtgttc cttccagaat ctccaccaga ggaaggctgg gaggagcaga tgtgtgtcaa
     1741 actctcggag aaccgtgccc ggaaaaaaag gctgaatatt attattgtgg ctgaaggagc
     1801 aattgatacc caaaataaac ccatcacctc tgagaaaatc aaagagcttg tcgtcacgca
     1861 gctgggctat gacacacgtg tgaccatcct cgggcacgtg cagagaggag ggaccccttc
     1921 ggcattcgac aggatcttgg ccagccgcat gggagtggag gcagtcatcg ccttgctaga
     1981 ggccaccccg gacaccccag cttgcgtcgt gtcactgaac gggaaccacg ccgtgcgcct
     2041 gccgctgatg gagtgcgtgc agatgactca ggatgtgcag aaggcgatgg acgagaggag
     2101 atttcaagat gcggttcgac tccgagggag gagctttgcg ggcaacctga acacctacaa
     2161 gcgacttgcc atcaagctgc cggatgatca gatcccaaag accaattgca acgtagctgt
     2221 catcaacgtg ggggcacccg cggctgggat gaacgcagcc gtacgctcag ctgtgcgcgt
     2281 gggcattgcc gacggccaca ggatgctcgc catctatgat ggctttgacg gcttcgccaa
     2341 gggccagatc aaagaaatcg gctggacaga tgtcgggggc tggaccggcc aaggaggctc
     2401 cattcttggg acaaaacgcg ttctcccggg gaagtacttg gaagagatcg ccacacagat
     2461 gcgcacgcac agcatcaacg cgctgctgat catcggtgga ttcgaggcct acctgggact
     2521 cctggagctg tcagccgccc gggagaagca cgaggagttc tgtgtcccca tggtcatggt
     2581 tcccgctact gtgtccaaca atgtgccggg ttccgatttc agcatcgggg cagacaccgc
     2641 cctgaacact atcaccgaca cctgcgaccg catcaagcag tccgccagcg gaaccaagcg
     2701 gcgcgtgttc atcatcgaga ccatgggcgg ctactgtggc tacctggcca acatgggggg
     2761 gctcgcggcc ggagctgatg ccgcatacat tttcgaagag cccttcgaca tcagggatct
     2821 gcagtccaac gtggagcacc tgacggagaa aatgaagacc accatccaga gaggccttgt
     2881 gctcagaaat gagagctgca gtgaaaacta caccaccgac ttcatttacc agctgtattc
     2941 agaagagggc aaaggcgtgt ttgactgcag gaagaacgtg ctgggtcaca tgcagcaggg
     3001 tggggcaccc tctccatttg atagaaactt tggaaccaaa atctctgcca gagctatgga
     3061 gtggatcact gcaaaactca aggaggcccg gggcagagga aaaaaattta ccaccgatga
     3121 ttccatttgt gtgctgggaa taagcaaaag aaacgttatt tttcaacctg tggcagagct
     3181 gaagaagcaa acggattttg agcacaggat tcccaaagaa cagtggtggc tcaagctacg
     3241 gcccctcatg aaaatcctgg ccaagtacaa ggccagctat gacgtgtcgg actcaggcca
     3301 gctggaacat gtgcagccct ggagtgtcgc ggccgcacat caccatcatc accattgact
     3361 cgagtctaga gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca
     3421 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     3481 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     3541 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     3601 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag
     3661 ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     3721 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     3781 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     3841 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     3901 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     3961 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     4021 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     4081 acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     4141 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     4201 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     4261 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     4321 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     4381 tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     4441 aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga
     4501 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     4561 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     4621 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     4681 aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     4741 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     4801 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     4861 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     4921 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     4981 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc
     5041 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     5101 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     5161 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     5221 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     5281 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     5341 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     5401 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     5461 agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     5521 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     5581 aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt
     5641 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     5701 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     5761 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     5821 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     5881 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     5941 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     6001 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     6061 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     6121 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     6181 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     6241 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     6301 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     6361 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     6421 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     6481 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     6541 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtttt tttgtttgca
     6601 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     6661 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     6721 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     6781 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     6841 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     6901 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     6961 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     7021 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     7081 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     7141 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     7201 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     7261 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     7321 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     7381 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     7441 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     7501 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     7561 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     7621 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     7681 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     7741 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     7801 tcgacggatc gggagatctc ccgatcccct atggt
//
