LOCUS       V022923                 7020 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V022923
VERSION     V022923
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..1694
                     /label="NEK6(NM_014397)"
                     /note="NEK6(NM_014397)"
                     /gene="NEK6"
     CDS             1707..1772
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    1796..2384
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    2450..2498
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      2700..3128
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        3142..3471
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             3538..4332
                     /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    4508..4629
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(4678..4694)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    4702..4718
                     /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(4726..4756)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    4771..4792
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(5080..5668)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(5839..6699)
                     /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(6700..6804)
                     /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 ccaccatggc aggacagccc ggccacatgc
      781 cccatggagg gagttccaac aacctctgcc acaccctggg gcctgtgcat cctcctgacc
      841 cacagaggca tcccaacacg ctgtcttttc gctgctcgct ggcggacttc cagatcgaaa
      901 agaagatagg ccgaggacag ttcagcgagg tgtacaaggc cacctgcctg ctggacagga
      961 agacagtggc tctgaagaag gtgcagatct ttgagatgat ggacgccaag gcgaggcagg
     1021 actgtgtcaa ggagatcggc ctcttgaagc aactgaacca cccaaatatc atcaagtatt
     1081 tggactcgtt tatcgaagac aacgagctga acattgtgct ggagttggct gacgcagggg
     1141 acctctcgca gatgatcaag tactttaaga agcagaagcg gctcatcccg gagaggacag
     1201 tatggaagta ctttgtgcag ctgtgcagcg ccgtggagca catgcattca cgccgggtga
     1261 tgcaccgaga catcaagcct gccaacgtgt tcatcacagc cacgggcgtc gtgaagctcg
     1321 gtgaccttgg tctgggccgc ttcttcagct ctgagaccac cgcagcccac tccctagtgg
     1381 ggacgcccta ctacatgtca ccggagagga tccatgagaa cggctacaac ttcaagtccg
     1441 acatctggtc cctgggctgt ctgctgtacg agatggcagc cctccagagc cccttctatg
     1501 gagataagat gaatctcttc tccctgtgcc agaagatcga gcagtgtgac taccccccac
     1561 tccccgggga gcactactcc gagaagttac gagaactggt cagcatgtgc atctgccctg
     1621 acccccacca gagacctgac atcggatacg tgcaccaggt ggccaagcag atgcacatct
     1681 ggatgtccag caccctcgag tctagagact acaaagacca tgacggtgat tataaagatc
     1741 atgacatcga ctacaaggat gacgatgaca agtagtgagg gcccgatatc tcgacaatca
     1801 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     1861 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc
     1921 tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc
     1981 cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg
     2041 gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc
     2101 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg
     2161 cactgacaat tccgtggtgt tgtcggggaa gctgacgtcc tttccatggc tgctcgcctg
     2221 tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc
     2281 agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct
     2341 tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcctggaa acgggggagg
     2401 ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa
     2461 agacagaata aaacgcacgg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca
     2521 gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc
     2581 ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt
     2641 cggggcggca ggccctgcca tagcagatct gcgcagctgg ggctctaggg ggtatcccca
     2701 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     2761 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     2821 gttcgccggc tttccccgtc aagctctaaa tcggggcatc cctttagggt tccgatttag
     2881 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     2941 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     3001 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     3061 agggattttg gggatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     3121 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     3181 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     3241 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     3301 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     3361 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     3421 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     3481 ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg
     3541 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     3601 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     3661 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     3721 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     3781 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     3841 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     3901 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     3961 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     4021 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     4081 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     4141 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4201 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4261 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     4321 gagttcttct gagcgggact ctggggttcg cgaaatgacc gaccaagcga cgcccaacct
     4381 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     4441 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     4501 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     4561 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     4621 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc
     4681 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     4741 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     4801 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     4861 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     4921 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     4981 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     5041 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     5101 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     5161 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     5221 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcaatgctc
     5281 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     5341 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     5401 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     5461 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     5521 gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     5581 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     5641 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     5701 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     5761 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     5821 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     5881 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     5941 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     6001 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     6061 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     6121 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     6181 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     6241 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     6301 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     6361 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     6421 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     6481 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     6541 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     6601 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     6661 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     6721 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     6781 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg
     6841 atcgggagat ctcccgatcc cctatggtcg actctcagta caatctgctc tgatgccgca
     6901 tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta gtgcgcgagc
     6961 aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg
//
