LOCUS       V032071                 7674 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V032071
VERSION     V032071
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..2354
                     /label="NETO1(NM_001201465)"
                     /note="NETO1(NM_001201465)"
                     /gene="NETO1"
     CDS             2361..2426
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    2450..3038
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    3104..3152
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      3354..3782
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        3796..4125
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             4192..4986
                     /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    5162..5283
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(5332..5348)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    5356..5372
                     /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(5380..5410)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    5425..5446
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(5734..6322)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(6493..7353)
                     /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(7354..7458)
                     /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 ctcgaattcg ccaccatgat ccatgggcgc agcgtgcttc
      781 acattgtagc aagtttaatc atcctccatt tgtctggggc aaccaagaaa ggaacagaaa
      841 agcaaaccac ctcagaaaca cagaagtcag tgcagtgtgg aacttggaca aaacatgcag
      901 agggaggtat ctttacctct cccaactatc ccagcaagta tccccctgac cgggaatgca
      961 tctacatcat agaagccgct ccaagacagt gcattgaact ttactttgat gaaaagtact
     1021 ctattgaacc gtcttgggag tgcaaatttg atcatattga agttcgagat ggaccttttg
     1081 gcttttctcc aataattgga cgtttctgtg gacaacaaaa tccacctgtc ataaaatcca
     1141 gtggaagatt tctatggatt aaattttttg ctgatggaga gctggaatct atgggatttt
     1201 cagctcgata caatttcaca cctgatcctg actttaagga ccttggagct ttgaaaccat
     1261 taccagcgtg tgagtttgag atgggcggtt ccgaaggaat tgtggagtct atacaaatta
     1321 tgaaggaagg caaagctact gctagcgagg ctgttgattg caagtggtac atccgagcac
     1381 ctccacggtc caagatttac ttacgattct tggactatga gatgcagaat tcaaatgagt
     1441 gcaagaggaa ttttgtggct gtgtatgatg gaagcagttc cgtggaggat ttgaaagcta
     1501 agttctgtag cactgtggct aatgatgtca tgctacgcac gggtcttggg gtgatccgca
     1561 tgtgggcaga tgagggcagt cgaaacagcc gatttcagat gctcttcaca tcctttcaag
     1621 aacctccttg tgaaggcaac acattcttct gccatagtaa catgtgtatt aataatactt
     1681 tggtctgcaa tggactccag aactgtgtgt atccttggga tgaaaatcac tgtaaagaga
     1741 agaggaaaac cagcctgctg gaccagctga ccaacaccag tgggactgtc attggcgtga
     1801 cttcctgcat cgtgatcatc ctcattatca tctctgtcat cgtacagatc aaacagcctc
     1861 gtaaaaagta tgtccaaagg aaatcagact ttgaccagac agttttccag gaggtatttg
     1921 aacctcctca ttatgagtta tgcactctca gagggacagg agctacagct gactttgcag
     1981 atgtggcaga tgactttgaa aattaccata aactgcggag gtcatcttcc aaatgcattc
     2041 atgaccatca ctgtggatca cagctgtcca gcactaaagg cagccgcagt aacctcagca
     2101 caagagatgc ttctatcttg acagagatgc ccacacagcc aggaaaaccc ctcatcccac
     2161 ccatgaacag aagaaatatc cttgtcatga aacacagcta ctcgcaagat gctgcagatg
     2221 cctgtgacat agatgaaatc gaagaggtgc cgaccaccag tcacaggctg tccagacacg
     2281 ataaagccgt ccagcggttc tgcctcattg ggtctctaag caaacatgaa tctgaataca
     2341 acacaactag ggtcggatcc gactacaaag accatgacgg tgattataaa gatcatgaca
     2401 tcgactacaa ggatgacgat gacaagtagt gagggcccga tatctcgaca atcaacctct
     2461 ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct
     2521 atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat
     2581 tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt
     2641 caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat
     2701 tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc
     2761 ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga
     2821 caattccgtg gtgttgtcgg ggaagctgac gtcctttcca tggctgctcg cctgtgttgc
     2881 cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga
     2941 ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc
     3001 tcagacgagt cggatctccc tttgggccgc ctccccgcct ggaaacgggg gaggctaact
     3061 gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag
     3121 aataaaacgc acgggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg
     3181 gcactctgtc gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt
     3241 ttccccaccc caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc
     3301 ggcaggccct gccatagcag atctgcgcag ctggggctct agggggtatc cccacgcgcc
     3361 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     3421 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     3481 cggctttccc cgtcaagctc taaatcgggg catcccttta gggttccgat ttagtgcttt
     3541 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     3601 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     3661 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     3721 tttggggatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     3781 ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
     3841 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     3901 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     3961 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     4021 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc
     4081 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct
     4141 tgtatatcca ttttcggatc tgatcaagag acaggatgag gatcgtttcg catgattgaa
     4201 caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac
     4261 tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg
     4321 cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag
     4381 gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt
     4441 gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg
     4501 tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg
     4561 catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga
     4621 gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag
     4681 gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat
     4741 ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt
     4801 tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg
     4861 gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt
     4921 tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc
     4981 ttctgagcgg gactctgggg ttcgcgaaat gaccgaccaa gcgacgccca acctgccatc
     5041 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     5101 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     5161 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     5221 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     5281 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     5341 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     5401 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     5461 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     5521 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     5581 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     5641 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     5701 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     5761 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     5821 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     5881 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcaat gctcacgctg
     5941 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     6001 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     6061 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     6121 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     6181 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     6241 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     6301 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     6361 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     6421 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     6481 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     6541 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6601 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     6661 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6721 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     6781 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     6841 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     6901 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     6961 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     7021 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     7081 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     7141 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     7201 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     7261 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     7321 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     7381 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     7441 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg
     7501 agatctcccg atcccctatg gtcgactctc agtacaatct gctctgatgc cgcatagtta
     7561 agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt
     7621 taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct tagg
//
