LOCUS       V024940                 6716 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V024940
VERSION     V024940
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..2925
                     /label="ARR3(NM_004312)"
                     /note="ARR3(NM_004312)"
                     /gene="ARR3"
     CDS             2932..2952
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /transl_table=1
                     /translation="ENLYFQS"
                     /label="TEV site"
     CDS             2953..3666
                     /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             3667..3690
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin®, an engineered
                     form of streptavidin"
                     /transl_table=1
                     /translation="WSHPQFEK"
                     /label="Strep-Tag II"
     CDS             3709..3732
                     /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    3804..3859
                     /note="rabbit&nbsp;β-globin polyadenylation signal (Gil and
                     Proudfoot, 1987)"
                     /label="β-globin poly(A) signal"
     primer_bind     complement(4219..4235)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    4243..4259
                     /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(4267..4297)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    4312..4333
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     promoter        4392..4587
                     /note="SV40 early promoter"
                     /label="SV40 promoter"
     polyA_signal    4593..4727
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      complement(4966..5554)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(5725..6585)
                     /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(6586..6690)
                     /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 gatgtccaag gtgtttaaga agaccagctc caatgggaag
     1801 ctctccatct acctggggaa acgggacttc gtggaccatg tggacacggt ggaacccatt
     1861 gacggtgttg tcctggttga tcctgagtac ttaaaatgtc gaaagttgtt tgtcatgttg
     1921 acatgtgcct ttcgctatgg ccgtgatgac ttggaagtga ttggtctgac gttccgaaaa
     1981 gatctgtatg tgcagaccct gcaagtggtc ccagctgaat ccagcagccc tcaggggccc
     2041 ctcacagtcc tacaggagcg actactgcac aagctagggg acaatgccta cccctttacc
     2101 ctgcagatgg tgaccaacct gccctgttct gtgacactgc agccaggtcc tgaagatgca
     2161 ggaaagccct gtgggattga ctttgaagtg aagagtttct gtgctgaaaa cccagaggag
     2221 acagtctcca agagagacta tgtgcggctg gttgtccgga aagtacaatt tgcaccaccg
     2281 gaggcaggcc ctggcccctc agcccagacc atccgccgct tccttctgtc agctcagccc
     2341 ctacaactcc aggcctggat ggacagggag gttcactacc acggagaacc catctctgtc
     2401 aatgtttcta tcaacaactg caccaacaag gtcatcaaaa aaatcaagat ttcagttgac
     2461 cagatcacag atgttgtcct gtattcacta gacaagtaca ccaagactgt gttcattcag
     2521 gaattcacgg agactgtagc tgctaattcc agcttctccc agagctttgc agtaacccca
     2581 atcctggctg ccagctgcca gaaacggggc ctggcactgg atggcaaact taagcatgaa
     2641 gataccaacc tggcctctag cacaattatt agaccgggaa tggacaaaga gctgctgggg
     2701 atcctggtgt cctacaaagt cagagtcaac ctgatggtgt cctgtggtgg catcctagga
     2761 gacctgacag ccagcgatgt tggtgtggag ctacccttgg tcctgatcca tccgaagcca
     2821 tctcatgagg ccgctagctc tgaggacata gtcatcgagg agtttacgcg gaaaggcgag
     2881 gaggagagcc agaaggctgt ggaggctgag ggagatgagg ggagcgaatt cgaaaacctg
     2941 tacttccaat ccgtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc
     3001 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat
     3061 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc
     3121 tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac
     3181 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc
     3241 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc
     3301 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc
     3361 ctggggcaca agctggagta caactacaac agccacaacg tctatatcat ggccgacaag
     3421 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg
     3481 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc
     3541 gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat
     3601 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg
     3661 tacaagtgga gccacccgca gttcgagaag ggcggcggca gcagcgcctg gagccacccg
     3721 cagttcgaga agtgaagatc tttttccctc tgccaaaaat tatggggaca tcatgaagcc
     3781 ccttgagcat ctgacttctg gctaataaag gaaatttatt ttcattgcaa tagtgtgttg
     3841 gaattttttg tgtctctcac tcggaaggac atatgggagg gcaaatcatt taaaacatca
     3901 gaatgagtat ttggtttaga gtttggcaac atatgcccat atgctggctg ccatgaacaa
     3961 aggtggctat aaagaggtca tcagtatatg aaacagcccc ctgctgtcca ttccttattc
     4021 catagaaaag ccttgacttg aggttagatt ttttttatat tttgttttgt gttatttttt
     4081 tctttaacat ccctaaaatt ttccttacat gttttactag ccagattttt cctcctctcc
     4141 tgactactcc cagtcatagc tgtccctctt ctcttatgaa gatccctcga cctgcagccc
     4201 aagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     4261 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     4321 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     4381 ccagcggatc cgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     4441 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     4501 atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc
     4561 ttttttggag gcctaggctt ttgcaaaaag ctaacttgtt tattgcagct tataatggtt
     4621 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     4681 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggatccgc tgcattaatg
     4741 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     4801 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     4861 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     4921 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     4981 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     5041 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     5101 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     5161 atgctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     5221 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     5281 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     5341 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     5401 tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     5461 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     5521 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     5581 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     5641 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     5701 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     5761 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     5821 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     5881 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     5941 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     6001 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     6061 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     6121 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     6181 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     6241 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     6301 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     6361 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     6421 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     6481 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     6541 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     6601 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     6661 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctg
//
