LOCUS       V018681                 8371 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V018681
VERSION     V018681
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             1015..3924
                     /label="PLK4(NM_014264)"
                     /note="PLK4(NM_014264)"
                     /gene="PLK4"
     polyA_signal    3971..4195
                     /note="bovine growth hormone polyadenylation signal"
                     /label="bGH poly(A) signal"
     rep_origin      4241..4669
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        4683..5012
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             5079..5873
                     /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    6047..6168
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(6217..6233)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6241..6257
                     /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(6265..6295)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6310..6331
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6619..7204)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7375..8235)
                     /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(8236..8340)
                     /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 ggtaccgagc tcggatccgc caccatggcg
     1021 acctgcatcg gggagaagat cgaggatttt aaagttggaa atctgcttgg taaaggatca
     1081 tttgctggtg tctacagagc tgagtccatt cacactggtt tggaagttgc aatcaaaatg
     1141 atagataaga aagccatgta caaagcagga atggtacaga gagtccaaaa tgaggtgaaa
     1201 atacattgcc aattgaaaca tccttctatc ttggagcttt ataactattt tgaagatagc
     1261 aattatgtgt atctggtatt agaaatgtgc cataatggag aaatgaacag gtatctaaag
     1321 aatagagtga aacccttctc agaaaatgaa gctcgacact tcatgcacca gatcatcaca
     1381 gggatgttgt atcttcattc tcatggtata ctacaccggg acctcacact ttctaacctc
     1441 ctactgactc gtaatatgaa catcaagatt gctgattttg ggctggcaac tcaactgaaa
     1501 atgccacatg aaaagcacta tacattatgt ggaactccta actacatttc accagaaatt
     1561 gccactcgaa gtgcacatgg ccttgaatct gatgtttggt ccctgggctg tatgttttat
     1621 acattactta tcgggagacc acccttcgac actgacacag tcaagaacac attaaataaa
     1681 gtagtattgg cagattatga aatgccatct tttttgtcaa tagaggccaa ggaccttatt
     1741 caccagttac ttcgtagaaa tccagcagat cgtttaagtc tgtcttcagt attggaccat
     1801 ccttttatgt cccgaaattc ttcaacaaaa agtaaagatt taggaactgt ggaagactca
     1861 attgatagtg ggcatgccac aatttctact gcaattacag cttcttccag taccagtata
     1921 agtggtagtt tatttgacaa aagaagactt ttgattggtc agccactccc aaataaaatg
     1981 actgtatttc caaagaataa aagttcaact gatttttctt cttcaggaga tggaaacagt
     2041 ttttatactc agtggggaaa tcaagaaacc agtaatagtg gaaggggaag agtaattcaa
     2101 gatgcagaag aaaggccaca ttctcgatac cttcgtagag cttattcctc tgatagatct
     2161 ggcacttcta atagtcagtc tcaagcaaaa acatatacaa tggaacgatg tcactcagca
     2221 gaaatgcttt cagtgtccaa aagatcagga ggaggtgaaa atgaagagag gtactcaccc
     2281 acagacaaca atgccaacat ttttaacttc tttaaagaaa agacatccag tagttctgga
     2341 tcttttgaaa gacctgataa caatcaagca ctctccaatc atctttgtcc aggaaaaact
     2401 ccttttccat ttgcagaccc gacacctcag accgaaaccg tacaacagtg gtttgggaat
     2461 ctgcaaataa atgctcattt aagaaaaact actgaatatg acagcatcag cccaaaccgg
     2521 gacttccagg gccatccaga tttgcagaag gacacatcaa aaaatgcctg gactgataca
     2581 aaagtcaaaa agaactctga tgcttctgat aatgcacatt ctgtaaaaca gcaaaatacc
     2641 atgaaatata tgactgcact tcacagtaaa cctgagataa tccaacaaga atgtgttttt
     2701 ggctcagatc ctctttctga acagagcaag actaggggta tggagccacc atggggttat
     2761 cagaatcgta cattaagaag cattacatct ccgttggttg ctcacaggtt aaaaccaatc
     2821 agacagaaaa ccaaaaaggc tgtggtgagc atacttgatt cagaggaggt gtgtgtggag
     2881 cttgtaaagg agtatgcatc tcaagaatat gtgaaagaag ttcttcagat atctagtgat
     2941 ggaaatacga tcactattta ttatccaaat ggtggtagag gttttcctct tgctgataga
     3001 ccaccctcac ctactgacaa catcagtagg tacagctttg acaatttacc agaaaaatac
     3061 tggcgaaaat atcaatatgc ttccaggttt gtacagcttg taagatctaa atctcccaaa
     3121 atcacttatt ttacaagata tgctaaatgc attttgatgg agaattctcc tggtgctgat
     3181 tttgaggttt ggttttatga tggggtaaaa atacacaaaa cagaagattt cattcaggtg
     3241 attgaaaaga cagggaagtc ttacacttta aaaagtgaaa gtgaagttaa tagcttgaaa
     3301 gaggagataa aaatgtatat ggaccatgct aatgagggtc atcgtatttg tttagcactg
     3361 gaatccataa tttcagaaga ggaaaggaaa actaggagtg ctcccttttt cccaataatc
     3421 ataggaagaa aacctggtag tactagttca cctaaggcct tatcacctcc tccttctgtg
     3481 gattcaaatt acccaacgag agagagagca tctttcaaca gaatggtcat gcatagtgct
     3541 gcttctccaa cacaggcacc aatccttaat ccctctatgg ttacaaatga aggacttggt
     3601 cttacaacta cagcttctgg aacagacatc tcttctaata gtctaaaaga ttgtcttcct
     3661 aaatcagcac aacttttgaa atctgttttt gtgaaaaatg ttggttgggc tacacagtta
     3721 actagtggag ctgtgtgggt tcagtttaat gatgggtccc agttggttgt gcaggcagga
     3781 gtgtcttcta tcagttatac ctcaccaaat ggtcaaacaa ctaggtatgg agaaaatgaa
     3841 aaattaccag actacatcaa acagaaatta cagtgtctgt cttccatcct tttgatgttt
     3901 tctaatccga ctcctaattt tcattgactc gagtctagag ggcccgttta aacccgctga
     3961 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     4021 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     4081 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
     4141 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
     4201 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
     4261 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     4321 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     4381 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     4441 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     4501 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     4561 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     4621 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
     4681 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     4741 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     4801 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     4861 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     4921 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     4981 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
     5041 tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     5101 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     5161 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     5221 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     5281 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     5341 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     5401 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     5461 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     5521 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     5581 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     5641 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     5701 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     5761 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     5821 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     5881 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     5941 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     6001 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     6061 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     6121 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     6181 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     6241 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     6301 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     6361 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     6421 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     6481 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     6541 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     6601 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     6661 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     6721 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     6781 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     6841 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     6901 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     6961 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     7021 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     7081 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     7141 caccgctggt agcggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     7201 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     7261 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     7321 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     7381 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     7441 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     7501 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     7561 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     7621 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     7681 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     7741 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     7801 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     7861 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     7921 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     7981 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     8041 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     8101 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     8161 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     8221 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     8281 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     8341 cacatttccc cgaaaagtgc cacctgacgt c
//
