LOCUS       V019062                 9360 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V019062
VERSION     V019062
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     promoter        50..1228
                     /note="strong constitutive promoter for human elongation
                     factor EF-1α"
                     /label="EF-1α promoter"
     CDS             1281..4031
                     /label="HK2(NM_000189)"
                     /note="HK2(NM_000189)"
                     /gene="HK2"
     CDS             4047..4112
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    4136..4724
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    4790..4838
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      5040..5468
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        5482..5811
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             5878..6672
                     /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    6848..6969
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(7018..7034)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    7042..7058
                     /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(7066..7096)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    7111..7132
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7420..8008)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8179..9039)
                     /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(9040..9144)
                     /gene="bla"
                     /label="AmpR promoter"
ORIGIN
        1 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg gctccggtgc
       61 ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg
      121 caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta
      181 ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt
      241 gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc
      301 ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacctg
      361 gctgcagtac gtgattcttg atcccgagct tcgggttgga agtgggtggg agagttcgag
      421 gccttgcgct taaggagccc cttcgcctcg tgcttgagtt gaggcctggc ctgggcgctg
      481 gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt tcgataagtc
      541 tctagccatt taaaattttt gatgacctgc tgcgacgctt tttttctggc aagatagtct
      601 tgtaaatgcg ggccaagatc tgcacactgg tatttcggtt tttggggccg cgggcggcga
      661 cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag cgcggccacc
      721 gagaatcgga cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg gcctcgcgcc
      781 gccgtgtatc gccccgccct gggcggcaag gctggcccgg tcggcaccag ttgcgtgagc
      841 ggaaagatgg ccgcttcccg gccctgctgc agggagctca aaatggagga cgcggcgctc
      901 gggagagcgg gcgggtgagt cacccacaca aaggaaaagg gcctttccgt cctcagccgt
      961 cgcttcatgt gactccacgg agtaccgggc gccgtccagg cacctcgatt agttctcgag
     1021 cttttggagt acgtcgtctt taggttgggg ggaggggttt tatgcgatgg agtttcccca
     1081 cactgagtgg gtggagactg aagttaggcc agcttggcac ttgatgtaat tctccttgga
     1141 atttgccctt tttgagtttg gatcttggtt cattctcaag cctcagacag tggttcaaag
     1201 tttttttctt ccatttcagg tgtcgtgagg aattagcttg gtactggcta gcgtttaaac
     1261 ttaagcttgg taccgccacc atgattgcct cgcatctgct tgcctacttc ttcacggagc
     1321 tcaaccatga ccaagtgcag aaggttgacc agtatctcta ccacatgcgc ctctctgatg
     1381 agaccctctt ggagatctct aagcggttcc gcaaggagat ggagaaaggg cttggagcca
     1441 ccactcaccc tactgcagca gtgaagatgc tgcccacctt tgtgaggtcc actccagatg
     1501 ggacagaaca cggagagttc ctggctctgg atcttggagg gaccaacttc cgtgtgcttt
     1561 gggtgaaagt aacggacaat gggctccaga aggtggagat ggagaatcag atctatgcca
     1621 tccctgagga catcatgcga ggcagtggca cccagctgtt tgaccacatt gccgaatgcc
     1681 tggctaactt catggataag ctacaaatca aagacaagaa gctcccactg ggttttacct
     1741 tctcgttccc ctgccaccag actaaactag acgagagttt cctggtctca tggaccaagg
     1801 gattcaagtc cagtggagtg gaaggcagag acgttgtggc tctgatccgg aaggccatcc
     1861 agaggagagg ggactttgat atcgacattg tggctgtggt gaatgacaca gttgggacca
     1921 tgatgacctg tggttatgat gaccacaact gtgagattgg tctcattgtg ggcacgggca
     1981 gcaacgcctg ctacatggaa gagatgcgcc acatcgacat ggtggaaggc gatgaggggc
     2041 ggatgtgtat caatatggag tggggggcct tcggggacga tggctcgctc aacgacattc
     2101 gcactgagtt tgaccaggag attgacatgg gctcactgaa cccgggaaag caactgtttg
     2161 agaagatgat cagtgggatg tacatggggg agctggtgag gcttatcctg gtgaagatgg
     2221 ccaaggagga gctgctcttt ggggggaagc tcagcccaga gcttctcaac accggtcgct
     2281 ttgagaccaa agacatctca gacattgaag gggagaagga tggcatccgg aaggcccgtg
     2341 aggtcctgat gcggttgggc ctggacccga ctcaggagga ctgcgtggcc actcaccgga
     2401 tctgccagat cgtgtccaca cgctccgcca gcctgtgcgc agccaccctg gccgccgtgc
     2461 tgcagcgcat caaggagaac aaaggcgagg agcggctgcg ctctactatt ggggtcgacg
     2521 gttccgtcta caagaaacac ccccattttg ccaagcgtct acataagacc gtgcggcggc
     2581 tggtgcccgg ctgcgatgtc cgcttcctcc gctccgagga tggcagtggc aaaggtgcag
     2641 ccatggtgac agcagtggct taccggctgg ccgatcaaca ccgtgcccgc cagaagacat
     2701 tagagcatct gcagctgagc catgaccagc tgctggaggt caagaggagg atgaaggtag
     2761 aaatggagcg aggtctgagc aaggagactc atgccagtgc ccccgtcaag atgctgccca
     2821 cctacgtgtg tgctaccccg gacggcacag agaaagggga cttcttggcc ttggaccttg
     2881 gaggaacaaa tttccgggtc ctgctggtcc gtgttcggaa tgggaagtgg ggtggagtgg
     2941 agatgcacaa caagatctac gccatcccgc aggaggtcat gcacggcacc ggggacgagc
     3001 tctttgacca cattgtccag tgcatcgcgg acttcctcga gtacatgggc atgaagggcg
     3061 tgtccctgcc tctgggtttt accttctcct tcccctgcca gcagaacagc ctggacgaga
     3121 gcatcctcct caagtggaca aaaggcttca aggcatctgg ctgcgagggc gaggacgtgg
     3181 tgaccctgct gaaggaagcg atccaccggc gagaggagtt tgacctggat gtggttgctg
     3241 tggtgaacga cacagtcgga actatgatga cctgtggctt tgaagaccct cactgtgaag
     3301 ttggcctcat tgttggcacg ggcagcaatg cctgctacat ggaggagatg cgcaacgtgg
     3361 aactggtgga aggagaagag gggcggatgt gtgtgaacat ggaatggggg gccttcgggg
     3421 acaatggatg cctagatgac ttccgcacag aatttgatgt ggctgtggat gagctttcac
     3481 tcaaccccgg caagcagagg ttcgagaaaa tgatcagtgg aatgtacctg ggtgagattg
     3541 tccgtaacat tctcatcgat ttcaccaagc gtggactact cttccgaggc cgcatctcag
     3601 agcggctcaa gacaaggggc atctttgaaa ccaagttctt gtctcagatt gagagtgact
     3661 gcctggccct gctgcaagtc cgagccatcc tgcaacactt agggcttgag agcacctgtg
     3721 acgacagcat cattgttaag gaggtgtgca ctgtggtggc ccggcgggca gcccagctct
     3781 gtggcgcagg catggccgct gtggtggaca ggatacgaga aaaccgtggg ctggacgctc
     3841 tcaaagtgac agtgggtgtg gatgggaccc tctacaagct acatcctcac tttgccaaag
     3901 tcatgcatga gacagtgaag gacctggctc cgaaatgtga tgtgtctttc ctgcagtcag
     3961 aggatggcag cgggaagggg gcggcgctca tcactgctgt ggcctgccgc atccgtgagg
     4021 ctggacagcg aatgcggccg ctcgaggact acaaagacca tgacggtgat tataaagatc
     4081 atgacatcga ctacaaggat gacgatgaca agtagtgagg gcccgatatc tcgacaatca
     4141 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     4201 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc
     4261 tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc
     4321 cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg
     4381 gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc
     4441 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg
     4501 cactgacaat tccgtggtgt tgtcggggaa gctgacgtcc tttccatggc tgctcgcctg
     4561 tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc
     4621 agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct
     4681 tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcctggaa acgggggagg
     4741 ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa
     4801 agacagaata aaacgcacgg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca
     4861 gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc
     4921 ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt
     4981 cggggcggca ggccctgcca tagcagatct gcgcagctgg ggctctaggg ggtatcccca
     5041 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     5101 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     5161 gttcgccggc tttccccgtc aagctctaaa tcggggcatc cctttagggt tccgatttag
     5221 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     5281 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     5341 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     5401 agggattttg gggatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     5461 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     5521 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     5581 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     5641 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     5701 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     5761 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     5821 ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg
     5881 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     5941 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     6001 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     6061 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     6121 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     6181 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     6241 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     6301 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     6361 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     6421 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     6481 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     6541 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     6601 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     6661 gagttcttct gagcgggact ctggggttcg cgaaatgacc gaccaagcga cgcccaacct
     6721 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     6781 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     6841 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     6901 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     6961 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc
     7021 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     7081 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     7141 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     7201 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     7261 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     7321 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     7381 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     7441 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     7501 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     7561 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcaatgctc
     7621 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     7681 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     7741 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     7801 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     7861 gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     7921 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     7981 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     8041 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     8101 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     8161 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     8221 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     8281 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     8341 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     8401 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     8461 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     8521 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     8581 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     8641 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     8701 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     8761 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     8821 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     8881 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     8941 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     9001 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     9061 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     9121 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg
     9181 atcgggagat ctcccgatcc cctatggtcg actctcagta caatctgctc tgatgccgca
     9241 tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta gtgcgcgagc
     9301 aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg
//
