LOCUS       V045213                 8588 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V045213
VERSION     V045213
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             771..2420
                     /codon_start=1
                     /note="synthetic luc2 version of the luciferase gene"
                     /product="firefly luciferase"
                     /protein_id=""
                     /transl_table=1
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
                     /label="luciferase"
     CDS             2424..2477
                     /codon_start=1
                     /note="Eukaryotic ribosomes fail to insert a peptide bond
                     between the Gly and Pro residues, yielding separate
                     polypeptides."
                     /product="2A peptide from&nbsp;Thosea asigna virus capsid
                     protein"
                     /protein_id=""
                     /transl_table=1
                     /translation="EGRGSLLTCGDVEENPGP"
                     /label="T2A"
     CDS             2484..3194
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from
                     mRuby2, with improved brightness and photostability (Bajar
                     et al., 2016)"
                     /transl_table=1
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD
                     IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV
                     AKYSNLGGGMDELYK"
                     /label="mRuby3"
     misc_feature    3364..3952
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    4018..4066
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      4268..4696
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        4710..5039
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             5106..5900
                     /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    6076..6197
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(6246..6262)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6270..6286
                     /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(6294..6324)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6339..6360
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6648..7236)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7407..8267)
                     /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(8268..8372)
                     /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 ctcgaattca cagcaggaaa ctgcaccagc atggaagatg
      781 ccaaaaacat taagaagggc ccagcgccat tctacccact cgaagacggg accgccggcg
      841 agcagctgca caaagccatg aagcgctacg ccctggtgcc cggcaccatc gcctttaccg
      901 acgcacatat cgaggtggac attacctacg ccgagtactt cgagatgagc gttcggctgg
      961 cagaagctat gaagcgctat gggctgaata caaaccatcg gatcgtggtg tgcagcgaga
     1021 atagcttgca gttcttcatg cccgtgttgg gtgccctgtt catcggtgtg gctgtggccc
     1081 cagctaacga catctacaac gagcgcgagc tgctgaacag catgggcatc agccagccca
     1141 ccgtcgtatt cgtgagcaag aaagggctgc aaaagatcct caacgtgcaa aagaagctac
     1201 cgatcataca aaagatcatc atcatggata gcaagaccga ctaccagggc ttccaaagca
     1261 tgtacacctt cgtgacttcc catttgccac ccggcttcaa cgagtacgac ttcgtgcccg
     1321 agagcttcga ccgggacaaa accatcgccc tgatcatgaa cagtagtggc agtaccggat
     1381 tgcccaaggg cgtagcccta ccgcaccgca ccgcttgtgt ccgattcagt catgcccgcg
     1441 accccatctt cggcaaccag atcatccccg acaccgctat cctcagcgtg gtgccatttc
     1501 accacggctt cggcatgttc accacgctgg gctacttgat ctgcggcttt cgggtcgtgc
     1561 tcatgtaccg cttcgaggag gagctattct tgcgcagctt gcaagactat aagattcaat
     1621 ctgccctgct ggtgcccaca ctatttagct tcttcgctaa gagcactctc atcgacaagt
     1681 acgacctaag caacttgcac gagatcgcca gcggcggggc gccgctcagc aaggaggtag
     1741 gtgaggccgt ggccaaacgc ttccacctac caggcatccg ccagggctac ggcctgacag
     1801 aaacaaccag cgccattctg atcacccccg aaggggacga caagcctggc gcagtaggca
     1861 aggtggtgcc cttcttcgag gctaaggtgg tggacttgga caccggtaag acactgggtg
     1921 tgaaccagcg cggcgagctg tgcgtccgtg gccccatgat catgagcggc tacgttaaca
     1981 accccgaggc tacaaacgct ctcatcgaca aggacggctg gctgcacagc ggcgacatcg
     2041 cctactggga cgaggacgag cacttcttca tcgtggaccg gctgaagagc ctgatcaaat
     2101 acaagggcta ccaggtagcc ccagccgaac tggagagcat cctgctgcaa caccccaaca
     2161 tcttcgacgc cggggtcgcc ggcctgcccg acgacgatgc cggcgagctg cccgccgcag
     2221 tcgtcgtgct ggaacacggt aaaaccatga ccgagaagga gatcgtggac tatgtggcca
     2281 gccaggttac aaccgccaag aagctgcgcg gtggtgttgt gttcgtggac gaggtgccta
     2341 aaggactgac cggcaagttg gacgcccgca agatccgcga gattctcatt aaggccaaga
     2401 agggcggcaa gatcgccgtg cttgagggca gaggaagtct gctaacatgc ggtgacgtgg
     2461 aggagaatcc cggccctgct agcatggtgt ctaagggcga agagctgatc aaggaaaata
     2521 tgcgtatgaa ggtggtcatg gaaggttcgg tcaacggcca ccaattcaaa tgcacaggtg
     2581 aaggagaagg cagaccgtac gagggaactc aaaccatgag gatcaaagtc atcgagggag
     2641 gacccctgcc atttgccttt gacattcttg ccacgtcgtt catgtatggc agccgtactt
     2701 ttatcaagta cccggccgac atccctgatt tctttaaaca gtcctttcct gagggtttta
     2761 cttgggaaag agttacgaga tacgaagatg gtggagtcgt caccgtcacg caggacacca
     2821 gccttgagga tggcgagctc gtctacaacg tcaaggtcag aggggtaaac tttccctcca
     2881 atggtcccgt gatgcagaag aagaccaagg gttgggagcc taatacagag atgatgtatc
     2941 cagcagatgg tggtctgaga ggatacactg acatcgcact gaaagttgat ggtggtggcc
     3001 atctgcactg caacttcgtg acaacttaca ggtcaaaaaa gaccgtcggg aacatcaaga
     3061 tgcccggtgt ccatgccgtt gatcaccgcc tggaaaggat cgaggagagt gacaatgaaa
     3121 cctacgtagt gcaacgcgaa gtggcagttg ccaaatacag caaccttggt ggtggcatgg
     3181 acgagctgta caagtgaact ccattctggg accatctcca ggagtccatg agaggctttc
     3241 ttctcctatg tcccaattct cagaactcag atgtggctag accaaccagt gggaaactgc
     3301 cccagcttct cccaccatag ggggccggac ccccatcacc agcctaggat ccgatatctc
     3361 gacaatcaac ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt
     3421 gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc
     3481 cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag
     3541 ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc
     3601 actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc
     3661 cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg
     3721 ctgttgggca ctgacaattc cgtggtgttg tcggggaagc tgacgtcctt tccatggctg
     3781 ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc
     3841 ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt
     3901 cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc gcctggaaac
     3961 gggggaggct aactgaaaca cggaaggaga caataccgga aggaacccgc gctatgacgg
     4021 caataaaaag acagaataaa acgcacgggt gttgggtcgt ttgttcataa acgcggggtt
     4081 cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc
     4141 cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca
     4201 gccaacgtcg gggcggcagg ccctgccata gcagatctgc gcagctgggg ctctaggggg
     4261 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     4321 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     4381 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc ggggcatccc tttagggttc
     4441 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     4501 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     4561 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     4621 gatttataag ggattttggg gatttcggcc tattggttaa aaaatgagct gatttaacaa
     4681 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     4741 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     4801 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     4861 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     4921 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     4981 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     5041 agctcccggg agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt
     5101 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     5161 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     5221 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     5281 aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     5341 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     5401 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
     5461 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     5521 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     5581 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc
     5641 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     5701 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
     5761 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     5821 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     5881 ttcttgacga gttcttctga gcgggactct ggggttcgcg aaatgaccga ccaagcgacg
     5941 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     6001 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     6061 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     6121 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     6181 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     6241 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     6301 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     6361 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     6421 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     6481 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     6541 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     6601 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     6661 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     6721 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     6781 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     6841 caatgctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     6901 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     6961 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     7021 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     7081 actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     7141 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     7201 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     7261 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     7321 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     7381 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     7441 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     7501 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     7561 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     7621 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     7681 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     7741 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     7801 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     7861 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     7921 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     7981 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     8041 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     8101 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     8161 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     8221 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     8281 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     8341 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     8401 gtcgacggat cgggagatct cccgatcccc tatggtcgac tctcagtaca atctgctctg
     8461 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
     8521 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
     8581 tgcttagg
//
