LOCUS       V019258                 6472 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V019258
VERSION     V019258
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"
     regulatory      643..652
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             652..717
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             748..1215
                     /label="TMIE(NM_147196)"
                     /note="TMIE(NM_147196)"
                     /gene="TMIE"
     misc_feature    1248..1836
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    1902..1950
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      2152..2580
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        2594..2923
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             2990..3784
                     /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    3960..4081
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(4130..4146)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    4154..4170
                     /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(4178..4208)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    4223..4244
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(4532..5120)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(5291..6151)
                     /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(6152..6256)
                     /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 gctctggcta gcgccaccat ggactacaaa
      661 gaccatgacg gtgattataa agatcatgac atcgactaca aggatgacga tgacaaggtc
      721 tacaagcttg gtaccgagct cggatccatg gcggggtggc cgggcgcggg tcccctctgc
      781 gtgctgggcg gcgccgcact cggggtgtgc ctcgcggggg ttgccgggca gctggtggag
      841 cccagcacgg ccccacccaa gcccaagccg cctccgctga ccaaggagac agtggtgttc
      901 tgggacatgc gcctgtggca cgtggtgggc atcttttcgc tcttcgtgtt gtccatcatc
      961 atcacgctgt gctgtgtctt caactgtcgt gtgccacgga cccggaagga gatcgaagcc
     1021 cggtacctgc agcgaaaggc agccaagatg tacacagaca agctggagac tgtgccaccc
     1081 ctcaatgagc tcacagaagt cccaggagag gataagaaga agaagaagaa gaagaagaag
     1141 gacagtgtgg acacagtggc catcaaagta gaggaggatg agaagaatga ggccaagaag
     1201 aagaaaggag agaaatgact cgagtctaga gggcccgata tctcgacaat caacctctgg
     1261 attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat
     1321 gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt
     1381 tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca
     1441 ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg
     1501 ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg
     1561 aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca
     1621 attccgtggt gttgtcgggg aagctgacgt cctttccatg gctgctcgcc tgtgttgcca
     1681 cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc
     1741 ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc
     1801 agacgagtcg gatctccctt tgggccgcct ccccgcctgg aaacggggga ggctaactga
     1861 aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa
     1921 taaaacgcac gggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc
     1981 actctgtcga taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt
     2041 ccccacccca ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg
     2101 caggccctgc catagcagat ctgcgcagct ggggctctag ggggtatccc cacgcgccct
     2161 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     2221 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     2281 gctttccccg tcaagctcta aatcggggca tccctttagg gttccgattt agtgctttac
     2341 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     2401 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     2461 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     2521 tggggatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     2581 aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     2641 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     2701 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     2761 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     2821 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca
     2881 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg
     2941 tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca
     3001 agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg
     3061 ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg
     3121 cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc
     3181 agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt
     3241 cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc
     3301 atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca
     3361 tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc
     3421 acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg
     3481 gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct
     3541 cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc
     3601 tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc
     3661 tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta
     3721 cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt
     3781 ctgagcggga ctctggggtt cgcgaaatga ccgaccaagc gacgcccaac ctgccatcac
     3841 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
     3901 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca
     3961 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     4021 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     4081 atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt
     4141 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     4201 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     4261 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     4321 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     4381 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4441 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     4501 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4561 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4621 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4681 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc tcacgctgta
     4741 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4801 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4861 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4921 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat
     4981 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     5041 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     5101 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     5161 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     5221 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     5281 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5341 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5401 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5461 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5521 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5581 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     5641 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5701 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5761 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5821 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5881 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     5941 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     6001 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     6061 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     6121 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     6181 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6241 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag
     6301 atctcccgat cccctatggt cgactctcag tacaatctgc tctgatgccg catagttaag
     6361 ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta
     6421 agctacaaca aggcaaggct tgaccgacaa ttgcatgaag aatctgctta gg
//
