LOCUS       V047451                 6853 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V047451
VERSION     V047451
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     enhancer        4..383
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        385..662
                     /label="chicken β-actin promoter"
     intron          663..1680
                     /note="chimera between introns from chicken β-actin and
                     rabbit β-globin"
                     /label="chimeric intron"
     regulatory      1729..1738
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             1738..1761
                     /codon_start=1
                     /product="FLAG® epitope tag, followed by an enterokinase
                     cleavage site"
                     /transl_table=1
                     /translation="DYKDDDDK"
                     /label="FLAG"
     CDS             1768..2853
                     /label="Sbk2(NM_001146329)"
                     /note="Sbk2(NM_001146329)"
                     /gene="Sbk2"
     CDS             2884..3840
                     /codon_start=1
                     /product="mutant of the E. coli biotin ligase BioID
                     engineered for improved catalytic efficiency (Branon et
                     al., 2018)"
                     /transl_table=1
                     /translation="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIPLLNAKQILGQLDGGSVAVLPVVDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGSRGRKWFSPFGANLYLSMFWRLKRGPAAIGLGPVIGIVMAEALRKLG
                     ADKVRVKWPNDLYLQDRKLAGILVELAGITGDAAQIVIGAGINVAMRRVEESVVNQGWI
                     TLQEAGINLDRNTLAATLIRELRAALELFEQEGLAPYLPRWEKLDNFINRPVKLIIGDK
                     EIFGISRGIDKQGALLLEQDGVIKPWMGGEISLRSAEK"
                     /label="TurboID"
     polyA_signal    3940..3995
                     /note="rabbit&nbsp;β-globin polyadenylation signal (Gil and
                     Proudfoot, 1987)"
                     /label="β-globin poly(A) signal"
     primer_bind     complement(4356..4372)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    4380..4396
                     /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(4404..4434)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    4449..4470
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     promoter        4529..4724
                     /note="SV40 early promoter"
                     /label="SV40 promoter"
     polyA_signal    4730..4864
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      complement(5102..5690)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(5861..6721)
                     /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(6722..6826)
                     /gene="bla"
                     /label="AmpR promoter"
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca
      421 tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag
      481 cgatgggggc gggggggggg ggggggcccc ccccaggcgg ggcggggcgg ggcgaggggc
      541 ggggcggggc gaggcggaaa ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt
      601 ttccttttat ggcgaggcgg cggcggcggc ggccctataa aaagcgaagc gcgcggcggg
      661 cgggagtcgt tgcgcgctgc cttccccccg tgccccgctc cgccgccgcc tcgcgccgcc
      721 cgccccggct ctgactgacc gcgttactcc cacaggtgag cgggcgggac ggcccttctc
      781 ctccgggctg taattagcgc ttggtttaat gacggcttgt ttcttttctg tggctgcgtg
      841 aaagccttga ggggctccgg gagggccctt tgtgcggggg gagcggctcg gggggtgcgt
      901 gcgtgtgtgt gtgcgtgggg agcgccgcgt gcggctccgc gctgcccggc ggctgtgagc
      961 gctgcgggcg cggcgcgggg ctttgtgcgc tccgcagtgt gcgcgagggg agcgcggccg
     1021 ggggcggtgc cccgcggtgc ggggggggct gcgaggggaa caaaggctgc gtgcggggtg
     1081 tgtgcgtggg ggggtgagca gggggtgtgg gcgcgtcggt cgggctgcaa ccccccctgc
     1141 acccccctcc ccgagttgct gagcacggcc cggcttcggg tgcggggctc cgtacggggc
     1201 gtggcgcggg gctcgccgtg ccgggcgggg ggtggcggca ggtgggggtg ccgggcgggg
     1261 cggggccgcc tcgggccggg gagggctcgg gggaaggggc gcggcggccc ccggagcgcc
     1321 ggcggctgtc gaggcgcggc gagccgcagc cattgccttt tatggtaatc gtgcgagagg
     1381 gcgcagggac ttcctttgtc ccaaatctgt gcggagccga aatctgggag gcgccgccgc
     1441 accccctcta gcgggcgcgg ggcgaagcgg tgcggcgccg gcaggaagga aatgggcggg
     1501 gagggccttc gtgcgtcgcc gcgccgccgt ccccttctcc ctctccagcc tcggggctgt
     1561 ccgcgggggg acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt
     1621 gtgaccggcg gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag
     1681 ctcctgggca acgtgctggt tgttgtgctg tctcatcatt ttggcaaagc caccatggat
     1741 tacaaggatg acgacgataa ggaattcatg ccaggcaaac agtcagaaga caagccaatg
     1801 gaagtgtcaa ctgtggagga tggaggtgat gaaggcctgg gaggcctaac ggtagaggag
     1861 ctgcagcagg ggcaggaggc tgctctggcc ttggaggaca tgatggctct gagtgcccag
     1921 accctggtcc aaactgaggt ggaagagctg tacgaggaag tgcgtcccct gggccagggc
     1981 cggttcggcc gtgtcctgct ggttactcac cgccagaaag gaacacccct ggcgctgaag
     2041 cagctcccga agcaatccac ttccctccgt ggcttcctgt atgagttctg cgtgggcctc
     2101 tcacttggca cacattctgc catcgtgaca gcctatggca ttggcattga gtcagccaat
     2161 tcctacagct tcctgacaga acctgtcctc catggggacc taatcacctt catccagccc
     2221 aaggtgggtc tcccacagcc agcagcccag cgctgtgcag cccaactggc ctctgccctg
     2281 gaacacattc actcgcatgg cctggtgtac cgggacttga agccagagaa tgtgctggtg
     2341 tgtgacccag cctgccagcg cgtcaagctg acagacttcg gccacacccg gccccgaggg
     2401 accctgctcc gactcactgg gccacctatc ccctacactg cccctgagct ctgtgcccca
     2461 ccacccctcc ccgagggact acccatccaa ccttccctgg atgcctgggc cctaggggtc
     2521 ctgatcttct gcctccttac tggctacttc ccctgggacc agcccctggt ggaggttgac
     2581 cctttctttg aggactttct catctggcag gcttctggtc agccccaaga ccggccacag
     2641 ccctggtata gcctgtcacc tgcagcagac actctcctat gggggctgct agatcctcac
     2701 ccccggaaga ggaaccctgt gggctccatc aagagctatc taggccaacc ctggaagcag
     2761 agggaagggg aggctgagga attggcaacg gagctgagag aagatgggtg gagaggaggg
     2821 caggaggctg caaaggggga gcaacctgca tgcggtggag gcggatcagg aggcggtgga
     2881 tctaaagaca atactgtgcc tctgaagctg atcgctctcc tggctaatgg cgagttccat
     2941 agtggcgaac agctgggaga aaccctgggc atgtccaggg ccgctatcaa caagcacatt
     3001 cagactctgc gcgactgggg cgtggacgtg ttcaccgtgc ccggaaaggg ctactctctg
     3061 cccgagccta tcccgctgct gaacgctaaa cagattctgg gacagctgga cggcgggagc
     3121 gtggcagtcc tgcctgtggt cgactccacc aatcagtacc tgctggatcg aatcggcgag
     3181 ctgaagagtg gggatgcttg cattgcagaa tatcagcagg cagggagagg aagcagaggg
     3241 aggaaatggt tctctccttt tggagctaac ctgtacctga gtatgttttg gcgcctgaag
     3301 cggggaccag cagcaatcgg cctgggcccg gtcatcggaa ttgtcatggc agaagcgctg
     3361 cgaaagctgg gagcagacaa ggtgcgagtc aaatggccca atgacctgta tctgcaggat
     3421 agaaagctgg caggcatcct ggtggagctg gccggaataa caggcgatgc tgcacagatc
     3481 gtcattggcg ccgggattaa cgtggctatg aggcgcgtgg aggaaagcgt ggtcaatcag
     3541 ggctggatca cactgcagga agcagggatt aacctggaca ggaatactct ggccgctacg
     3601 ctgatccgag agctgcgggc agccctggaa ctgttcgagc aggaaggcct ggctccatat
     3661 ctgccacggt gggagaagct ggataacttc atcaatagac ccgtgaagct gatcattggg
     3721 gacaaagaga ttttcgggat tagccggggg attgataaac agggagccct gctgctggaa
     3781 caggacggag ttatcaaacc ctggatgggc ggagaaatca gtctgcggtc tgccgaaaag
     3841 tgactcgagc tagcagatct taattaatta aggatctttt tccctctgcc aaaaattatg
     3901 gggacatcat gaagcccctt gagcatctga cttctggcta ataaaggaaa tttattttca
     3961 ttgcaatagt gtgttggaat tttttgtgtc tctcactcgg aaggacatat gggagggcaa
     4021 atcatttaaa acatcagaat gagtatttgg tttagagttt ggcaacatat gcccatatgc
     4081 tggctgccat gaacaaaggt tggctataaa gaggtcatca gtatatgaaa cagccccctg
     4141 ctgtccattc cttattccat agaaaagcct tgacttgagg ttagattttt tttatatttt
     4201 gttttgtgtt atttttttct ttaacatccc taaaattttc cttacatgtt ttactagcca
     4261 gatttttcct cctctcctga ctactcccag tcatagctgt ccctcttctc ttatggagat
     4321 ccctcgacct gcagcccaag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat
     4381 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg
     4441 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag
     4501 tcgggaaacc tgtcgtgcca gcggatccgc atctcaatta gtcagcaacc atagtcccgc
     4561 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     4621 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
     4681 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagcta acttgtttat
     4741 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     4801 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     4861 gatccgctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     4921 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     4981 tcagctcact caaaggcggt aatacggtta tccacagaat cagggataac gcaggaaaga
     5041 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     5101 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     5161 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     5221 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     5281 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     5341 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     5401 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     5461 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     5521 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     5581 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     5641 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     5701 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     5761 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     5821 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     5881 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     5941 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     6001 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     6061 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     6121 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     6181 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     6241 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     6301 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     6361 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     6421 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     6481 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     6541 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     6601 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     6661 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     6721 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     6781 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     6841 aagtgccacc tgg
//
