LOCUS       V033336                 6543 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V033336
VERSION     V033336
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..1223
                     /label="IER3(NM_003897)"
                     /note="IER3(NM_003897)"
                     /gene="IER3"
     CDS             1230..1295
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    1319..1907
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    1973..2021
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      2223..2651
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        2665..2994
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             3061..3855
                     /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    4031..4152
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(4201..4217)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    4225..4241
                     /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(4249..4279)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    4294..4315
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(4603..5191)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(5362..6222)
                     /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(6223..6327)
                     /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 ccaccatgtg tcactctcgc agctgccacc
      781 cgaccatgac catcctgcag gccccgaccc cggccccctc caccatcccg ggaccccggc
      841 ggggctccgg tcctgagatc ttcaccttcg accctctccc ggagcccgca gcggcccctg
      901 ccgggcgccc cagcgcctct cgcgggcacc gaaagcgcag ccgcagggtt ctctaccctc
      961 gagtggtccg gcgccagctg ccagtcgagg aaccgaaccc agccaaaagg cttctctttc
     1021 tgctgctcac catcgtcttc tgccagatcc tgatggctga agagggtgtg ccggcgcccc
     1081 tgcctccaga ggacgcccct aacgccgcat ccctggcgcc cacccctgtg tccgccgtcc
     1141 tcgagccctt taatctgact tcggagccct cggactacgc tctggacctc agcactttcc
     1201 tccagcaaca cccggccgcc ttcggatccg actacaaaga ccatgacggt gattataaag
     1261 atcatgacat cgactacaag gatgacgatg acaagtagtg agggcccgat atctcgacaa
     1321 tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc
     1381 ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat
     1441 ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg
     1501 gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg
     1561 ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat
     1621 tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt
     1681 gggcactgac aattccgtgg tgttgtcggg gaagctgacg tcctttccat ggctgctcgc
     1741 ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa
     1801 tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg
     1861 ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcctg gaaacggggg
     1921 aggctaactg aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata
     1981 aaaagacaga ataaaacgca cgggtgttgg gtcgtttgtt cataaacgcg gggttcggtc
     2041 ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt
     2101 ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa
     2161 cgtcggggcg gcaggccctg ccatagcaga tctgcgcagc tggggctcta gggggtatcc
     2221 ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     2281 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     2341 cacgttcgcc ggctttcccc gtcaagctct aaatcggggc atccctttag ggttccgatt
     2401 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     2461 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     2521 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     2581 ataagggatt ttggggattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     2641 taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     2701 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     2761 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     2821 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     2881 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct
     2941 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc
     3001 ccgggagctt gtatatccat tttcggatct gatcaagaga caggatgagg atcgtttcgc
     3061 atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc
     3121 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca
     3181 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg
     3241 caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg
     3301 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag
     3361 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg
     3421 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc
     3481 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa
     3541 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac
     3601 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat
     3661 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac
     3721 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc
     3781 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt
     3841 gacgagttct tctgagcggg actctggggt tcgcgaaatg accgaccaag cgacgcccaa
     3901 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     3961 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     4021 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     4081 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     4141 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
     4201 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     4261 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     4321 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     4381 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     4441 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     4501 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     4561 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     4621 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     4681 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     4741 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcaatg
     4801 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     4861 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     4921 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     4981 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     5041 aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     5101 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     5161 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     5221 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     5281 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     5341 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     5401 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     5461 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     5521 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     5581 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     5641 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     5701 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     5761 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     5821 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     5881 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     5941 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     6001 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     6061 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     6121 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     6181 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     6241 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     6301 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcga
     6361 cggatcggga gatctcccga tcccctatgg tcgactctca gtacaatctg ctctgatgcc
     6421 gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg
     6481 agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt
     6541 agg
//
