LOCUS       V032259                 7216 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V032259
VERSION     V032259
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     enhancer        235..614
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        615..818
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     promoter        863..881
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /label="T7 promoter"
     regulatory      913..922
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             922..987
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             991..2736
                     /label="Pxk(NM_145458)"
                     /note="Pxk(NM_145458)"
                     /gene="Pxk"
     polyA_signal    2816..3040
                     /note="bovine growth hormone polyadenylation signal"
                     /label="bGH poly(A) signal"
     rep_origin      3086..3514
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        3528..3857
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             3924..4718
                     /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    4892..5013
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(5062..5078)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    5086..5102
                     /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(5110..5140)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    5155..5176
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(5464..6049)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(6220..7080)
                     /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(7081..7185)
                     /gene="bla"
                     /label="AmpR promoter"
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagccaccat ggactacaaa gaccatgacg gtgattataa agatcatgac
      961 atcgactaca aggatgacga tgacaagctt atggccttca tggagaagcc gccggctggc
     1021 aaggtgctgc tggacgacac ggtgccgctg acagcggccg tcgaggcgag ccagagtctg
     1081 cagtcgcaca cggaatatat catccgggtg caaagaggaa tttctgcaga aaacagctgg
     1141 cagatcgtca gaagatacag tgacttcgat ctgcttaaca acagcttgca aattacaggc
     1201 ctgagcctgc ctcttcctcc taagaaactg attggaaaca tggaccgtga gttcatcgct
     1261 gagaggcagc gaggcctgca gaactacctt aacgtcatca tggcaaatca tgtcttgtcc
     1321 aactgtgagc tgcttaagaa gttcctagac ccaaacaact attctgcaaa ctatactgag
     1381 atcgccttac agcaggtttc catgttcttc cgatcagaac caaagtggga ggtggtagaa
     1441 ccactgaagg acataggttg gaggataagg aagaaatatt tcttgatgaa aataaaaaac
     1501 caaccaaagg agcggctggt gctaagctgg gctgatcttg gcccagacaa gtacttgtca
     1561 gataaagatt ttcagtgtct aatcaaactt ctgccttcct gtgtgcaccc ttacatctat
     1621 cgggtcacct ttgccacggc tagcgagtcc tccgcattgc tgattagagc ctttaatgag
     1681 aagggaactt tgaaggatct gatctacaag gcaaaaccaa aggacccctt cctaaagaaa
     1741 tactgcaacc ctaagaaaac ccagggcctt gaactccagc aaataaaaac atatgggcgg
     1801 cagatattag aggcactaaa gtttcttcat gacaagggat tcccttacgg ccacctccat
     1861 gcagccaatg tgatgctgga tggcaacact tgccggctgc tagacctcga aaactcactg
     1921 ctgggtctgc cttccttcta ccgatcctac ttcacacaat tcaggaaaat caatacattg
     1981 gagagtgtag atgtgcactg ttttggccac ttactctatg aaatgactta tggacgacct
     2041 cccgactccg tgcctgtgga ctcctttcct cctgcatcat ccttggctgt ggtggccgta
     2101 ttggagtcta cgctgtcttg tgaagcctgt aaaaatggca tgcctaccgt ctcccggctc
     2161 ttacagatgc cattattcag tgatgtttta ctaacaactt ctgaaaagcc acagtttaag
     2221 attcccacaa aactaaaaga ggcattgcga attgccaagg aatgcataga gaagagactc
     2281 accgaggaac agaagcagat tcaccagcat cgaagactaa cgagagccca gtcacaccat
     2341 gggtctgagg aagaaaggaa aaggagaaag atcttagctc gaaagaagtc aaaacgatct
     2401 gctgttgaaa acagtgagga gcagccagtg aagcacagta actccaataa ctcagcagga
     2461 tctggggcca gctcacctct cacatcccca tcatctccaa ctccaccctc cacagcagga
     2521 ctgtcttcag cattacctcc acctcctccg ccacctcctc ctccacctcc accagcaggt
     2581 ccctcgccaa cctcagccac ggagatgcct gctccgttcc tgccccagcc tgtgaatggt
     2641 gtgaaccgtg gggccttgct cagttccatc cagaatttcc aaaagggaac tttgcggaaa
     2701 gcccaaacct gtgatcacag cgctcccaag atcggctgag aattctgcag atatccagca
     2761 cagtggcggc cgctcgagtc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg
     2821 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     2881 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     2941 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     3001 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc
     3061 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt
     3121 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     3181 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     3241 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     3301 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     3361 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     3421 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     3481 aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag
     3541 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     3601 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     3661 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     3721 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     3781 aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     3841 gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag
     3901 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     3961 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     4021 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     4081 tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga
     4141 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     4201 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     4261 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     4321 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     4381 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     4441 ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     4501 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     4561 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     4621 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     4681 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     4741 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     4801 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     4861 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     4921 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     4981 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     5041 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     5101 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     5161 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     5221 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     5281 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     5341 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     5401 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     5461 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     5521 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     5581 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     5641 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     5701 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     5761 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     5821 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     5881 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     5941 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     6001 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     6061 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     6121 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     6181 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     6241 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     6301 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     6361 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     6421 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     6481 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     6541 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     6601 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     6661 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     6721 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     6781 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     6841 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     6901 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     6961 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     7021 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     7081 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     7141 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     7201 agtgccacct gacgtc
//
