LOCUS       V025088                 7112 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V025088
VERSION     V025088
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     enhancer        4..383
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        386..661
                     /label="chicken β-actin promoter"
     intron          662..1674
                     /note="chimera between introns from chicken β-actin and
                     rabbit β-globin"
                     /label="chimeric intron"
     regulatory      1681..1690
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             1690..1755
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             1762..3321
                     /label="ADRA1B(NM_000679)"
                     /note="ADRA1B(NM_000679)"
                     /gene="ADRA1B"
     CDS             3328..3348
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /transl_table=1
                     /translation="ENLYFQS"
                     /label="TEV site"
     CDS             3349..4062
                     /codon_start=1
                     /note="mammalian codon-optimized"
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /transl_table=1
                     /translation="V,SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
                     /label="EGFP"
     CDS             4063..4086
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin®, an engineered
                     form of streptavidin"
                     /transl_table=1
                     /translation="WSHPQFEK"
                     /label="Strep-Tag II"
     CDS             4105..4128
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin®, an engineered
                     form of streptavidin"
                     /transl_table=1
                     /translation="WSHPQFEK"
                     /label="Strep-Tag II"
     polyA_signal    4200..4255
                     /note="rabbit&nbsp;β-globin polyadenylation signal (Gil and
                     Proudfoot, 1987)"
                     /label="β-globin poly(A) signal"
     primer_bind     complement(4615..4631)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    4639..4655
                     /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(4663..4693)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    4708..4729
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     promoter        4788..4983
                     /note="SV40 early promoter"
                     /label="SV40 promoter"
     polyA_signal    4989..5123
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      complement(5362..5950)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(6121..6981)
                     /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(6982..7086)
                     /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 tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gcccccggag cgccggcggc
     1321 tgtcgaggcg cggcgagccg cagccattgc cttttatggt aatcgtgcga gagggcgcag
     1381 ggacttcctt tgtcccaaat ctgtgcggag ccgaaatctg ggaggcgccg ccgcaccccc
     1441 tctagcgggc gcggggcgaa gcggtgcggc gccggcagga aggaaatggg cggggagggc
     1501 cttcgtgcgt cgccgcgccg ccgtcccctt ctccctctcc agcctcgggg ctgtccgcgg
     1561 ggggacggct gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc
     1621 ggcggctcta gtgcctctgc taaccatgtt catgccttct tctttttcct acagctcctg
     1681 gccaccatgg actacaaaga ccatgacggt gattataaag atcatgacat cgactacaag
     1741 gatgacgatg acaagctcga gatgaatccc gacctggaca ccggccacaa cacatcagca
     1801 cctgcccact ggggagagtt gaaaaatgcc aacttcactg gccccaacca gacctcgagc
     1861 aactccacac tgccccagct ggacatcacc agggccatct ctgtgggcct ggtgctgggc
     1921 gccttcatcc tctttgccat cgtgggcaac atcctagtca tcttgtctgt ggcctgcaac
     1981 cggcacctgc ggacgcccac caactacttc attgtcaacc tggccatggc cgacctgctg
     2041 ttgagcttca ccgtcctgcc cttctcagcg gccctagagg tgctcggcta ctgggtgctg
     2101 gggcggatct tctgtgacat ctgggcagcc gtggatgtcc tgtgctgcac agcgtccatt
     2161 ctgagcctgt gcgccatctc catcgatcgc tacatcgggg tgcgctactc tctgcagtat
     2221 cccacgctgg tcacccggag gaaggccatc ttggcgctgc tcagtgtctg ggtcttgtcc
     2281 accgtcatct ccatcgggcc tctccttggg tggaaggagc cggcacccaa cgatgacaag
     2341 gagtgcgggg tcaccgaaga acccttctat gccctcttct cctctctggg ctccttctac
     2401 atccctctgg cggtcattct agtcatgtac tgccgtgtct atatagtggc caagagaacc
     2461 accaagaacc tagaggcagg agtcatgaag gagatgtcca actccaagga gctgaccctg
     2521 aggatccatt ccaagaactt tcacgaggac acccttagca gtaccaaggc caagggccac
     2581 aaccccagga gttccatagc tgtcaaactt tttaagttct ccagggaaaa gaaagcagct
     2641 aagacgttgg gcattgtggt cggtatgttc atcttgtgct ggctaccctt cttcatcgct
     2701 ctaccgcttg gctccttgtt ctccaccctg aagccccccg acgccgtgtt caaggtggtg
     2761 ttctggctgg gctacttcaa cagctgcctc aaccccatca tctacccatg ctccagcaag
     2821 gagttcaagc gcgctttcgt gcgcatcctc gggtgccagt gccgcggccg cggccgccgc
     2881 cgacgccgcc gccgccgtcg cctgggcggc tgcgcctaca cctaccggcc gtggacgcgc
     2941 ggcggctcgc tggagcgctc gcagtcgcgc aaggactcgc tggacgacag cggcagctgc
     3001 ctgagcggca gccagcggac cctgccctcg gcctcgccga gcccgggcta cctgggccgc
     3061 ggcgcgccac cgccagtcga gctgtgcgcc ttccccgagt ggaaggcgcc cggcgccctc
     3121 ctgagcctgc ccgcgcctga gccccccggc cgccgcggcc gccacgactc gggcccgctc
     3181 ttcaccttca agctcctgac cgagcccgag agccccggga ccgacggcgg cgccagcaac
     3241 ggaggctgcg aggccgcggc cgacgtggcc aacgggcagc cgggcttcaa aagcaacatg
     3301 cccctggcgc ccgggcagtt tgaattcgaa aacctgtact tccaatccgt gagcaagggc
     3361 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     3421 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
     3481 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg
     3541 acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc
     3601 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc
     3661 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
     3721 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     3781 tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac
     3841 ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag
     3901 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag
     3961 tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
     4021 accgccgccg ggatcactct cggcatggac gagctgtaca agtggagcca cccgcagttc
     4081 gagaagggcg gcggcagcag cgcctggagc cacccgcagt tcgagaagtg aagatctttt
     4141 tccctctgcc aaaaattatg gggacatcat gaagcccctt gagcatctga cttctggcta
     4201 ataaaggaaa tttattttca ttgcaatagt gtgttggaat tttttgtgtc tctcactcgg
     4261 aaggacatat gggagggcaa atcatttaaa acatcagaat gagtatttgg tttagagttt
     4321 ggcaacatat gcccatatgc tggctgccat gaacaaaggt ggctataaag aggtcatcag
     4381 tatatgaaac agccccctgc tgtccattcc ttattccata gaaaagcctt gacttgaggt
     4441 tagatttttt ttatattttg ttttgtgtta tttttttctt taacatccct aaaattttcc
     4501 ttacatgttt tactagccag atttttcctc ctctcctgac tactcccagt catagctgtc
     4561 cctcttctct tatgaagatc cctcgacctg cagcccaagc ttggcgtaat catggtcata
     4621 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     4681 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     4741 ctcactgccc gctttccagt cgggaaacct gtcgtgccag cggatccgca tctcaattag
     4801 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     4861 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     4921 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     4981 aaaaagctaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     5041 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     5101 atgtatctta tcatgtctgg atccgctgca ttaatgaatc ggccaacgcg cggggagagg
     5161 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     5221 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5281 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     5341 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     5401 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     5461 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     5521 cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc tcacgctgta ggtatctcag
     5581 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     5641 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     5701 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     5761 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     5821 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     5881 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     5941 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     6001 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     6061 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     6121 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     6181 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     6241 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     6301 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     6361 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     6421 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     6481 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     6541 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     6601 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     6661 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     6721 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     6781 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     6841 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     6901 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     6961 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     7021 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     7081 tccgcgcaca tttccccgaa aagtgccacc tg
//
