LOCUS       V032374                 8990 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V032374
VERSION     V032374
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     enhancer        202..581
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        582..785
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     promoter        830..848
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /label="T7 promoter"
     CDS             908..3691
                     /label="RB1(NM_000321)"
                     /note="RB1(NM_000321)"
                     /gene="RB1"
     CDS             3728..4444
                     /codon_start=1
                     /note="mammalian codon-optimized"
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /transl_table=1
                     /translation="M,V,SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKL
                     TLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFK
                     DDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNG
                     IKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMV
                     LLEFVTAAGITLGMDELYK"
                     /label="EGFP"
     CDS             4445..4468
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin®, an engineered
                     form of streptavidin"
                     /transl_table=1
                     /translation="WSHPQFEK"
                     /label="Strep-Tag II"
     CDS             4487..4510
                     /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    4557..4781
                     /note="bovine growth hormone polyadenylation signal"
                     /label="bGH poly(A) signal"
     rep_origin      4827..5255
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        5269..5598
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             5665..6459
                     /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    6633..6754
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(6803..6819)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6827..6843
                     /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(6851..6881)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6896..6917
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7205..7790)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7961..8821)
                     /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(8822..8926)
                     /gene="bla"
                     /label="AmpR promoter"
ORIGIN
        1 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
       61 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
      121 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
      181 tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta
      241 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
      301 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
      361 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
      421 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
      481 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
      541 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
      601 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
      661 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
      721 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
      781 gagctctctg gctaactaga gaacccactg cttactggct tatcgaaatt aatacgactc
      841 actataggga gacccaagct ggctagcgtt taaacttaag cttggtaccg agctcggatc
      901 cgccaccatg ccgcccaaaa ccccccgaaa aacggccgcc accgccgccg ctgccgccgc
      961 ggaacccccg gcaccgccgc cgccgccccc tcctgaggag gacccagagc aggacagcgg
     1021 cccggaggac ctgcctctcg tcaggcttga gtttgaagaa acagaagaac ctgattttac
     1081 tgcattatgt cagaaattaa agataccaga tcatgtcaga gagagagctt ggttaacttg
     1141 ggagaaagtt tcatctgtgg atggagtatt gggaggttat attcaaaaga aaaaggaact
     1201 gtggggaatc tgtatcttta ttgcagcagt tgacctagat gagatgtcgt tcacttttac
     1261 tgagctacag aaaaacatag aaatcagtgt ccataaattc tttaacttac taaaagaaat
     1321 tgataccagt accaaagttg ataatgctat gtcaagactg ttgaagaagt atgatgtatt
     1381 gtttgcactc ttcagcaaat tggaaaggac atgtgaactt atatatttga cacaacccag
     1441 cagttcgata tctactgaaa taaattctgc attggtgcta aaagtttctt ggatcacatt
     1501 tttattagct aaaggggaag tattacaaat ggaagatgat ctggtgattt catttcagtt
     1561 aatgctatgt gtccttgact attttattaa actctcacct cccatgttgc tcaaagaacc
     1621 atataaaaca gctgttatac ccattaatgg ttcacctcga acacccaggc gaggtcagaa
     1681 caggagtgca cggatagcaa aacaactaga aaatgataca agaattattg aagttctctg
     1741 taaagaacat gaatgtaata tagatgaggt gaaaaatgtt tatttcaaaa attttatacc
     1801 ttttatgaat tctcttggac ttgtaacatc taatggactt ccagaggttg aaaatctttc
     1861 taaacgatac gaagaaattt atcttaaaaa taaagatcta gatgcaagat tatttttgga
     1921 tcatgataaa actcttcaga ctgattctat agacagtttt gaaacacaga gaacaccacg
     1981 aaaaagtaac cttgatgaag aggtgaatgt aattcctcca cacactccag ttaggactgt
     2041 tatgaacact atccaacaat taatgatgat tttaaattca gcaagtgatc aaccttcaga
     2101 aaatctgatt tcctatttta acaactgcac agtgaatcca aaagaaagta tactgaaaag
     2161 agtgaaggat ataggataca tctttaaaga gaaatttgct aaagctgtgg gacagggttg
     2221 tgtcgaaatt ggatcacagc gatacaaact tggagttcgc ttgtattacc gagtaatgga
     2281 atccatgctt aaatcagaag aagaacgatt atccattcaa aattttagca aacttctgaa
     2341 tgacaacatt tttcatatgt ctttattggc gtgcgctctt gaggttgtaa tggccacata
     2401 tagcagaagt acatctcaga atcttgattc tggaacagat ttgtctttcc catggattct
     2461 gaatgtgctt aatttaaaag cctttgattt ttacaaagtg atcgaaagtt ttatcaaagc
     2521 agaaggcaac ttgacaagag aaatgataaa acatttagaa cgatgtgaac atcgaatcat
     2581 ggaatccctt gcatggctct cagattcacc tttatttgat cttattaaac aatcaaagga
     2641 ccgagaagga ccaactgatc accttgaatc tgcttgtcct cttaatcttc ctctccagaa
     2701 taatcacact gcagcagata tgtatctttc tcctgtaaga tctccaaaga aaaaaggttc
     2761 aactacgcgt gtaaattcta ctgcaaatgc agagacacaa gcaacctcag ccttccagac
     2821 ccagaagcca ttgaaatcta cctctctttc actgttttat aaaaaagtgt atcggctagc
     2881 ctatctccgg ctaaatacac tttgtgaacg ccttctgtct gagcacccag aattagaaca
     2941 tatcatctgg acccttttcc agcacaccct gcagaatgag tatgaactca tgagagacag
     3001 gcatttggac caaattatga tgtgttccat gtatggcata tgcaaagtga agaatataga
     3061 ccttaaattc aaaatcattg taacagcata caaggatctt cctcatgctg ttcaggagac
     3121 attcaaacgt gttttgatca aagaagagga gtatgattct attatagtat tctataactc
     3181 ggtcttcatg cagagactga aaacaaatat tttgcagtat gcttccacca ggccccctac
     3241 cttgtcacca atacctcaca ttcctcgaag cccttacaag tttcctagtt cacccttacg
     3301 gattcctgga gggaacatct atatttcacc cctgaagagt ccatataaaa tttcagaagg
     3361 tctgccaaca ccaacaaaaa tgactccaag atcaagaatc ttagtatcaa ttggtgaatc
     3421 attcgggact tctgagaagt tccagaaaat aaatcagatg gtatgtaaca gcgaccgtgt
     3481 gctcaaaaga agtgctgaag gaagcaaccc tcctaaacca ctgaaaaaac tacgctttga
     3541 tattgaagga tcagatgaag cagatggaag taaacatctc ccaggagagt ccaaatttca
     3601 gcagaaactg gcagaaatga cttctactcg aacacgaatg caaaagcaga aaatgaatga
     3661 tagcatggat acctcaaaca aggaagagaa agaattcggt ggaggcggat caggaggcgg
     3721 tggatctatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     3781 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     3841 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
     3901 gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca
     3961 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     4021 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     4081 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     4141 ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca
     4201 gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca
     4261 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     4321 caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca
     4381 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
     4441 caagtggagc cacccgcagt tcgagaaggg cggcggcagc agcgcctgga gccacccgca
     4501 gttcgagaag tgactcgagt ctagagggcc cgtttaaacc cgctgatcag cctcgactgt
     4561 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     4621 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     4681 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     4741 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     4801 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     4861 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     4921 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     4981 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     5041 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     5101 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     5161 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     5221 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
     5281 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     5341 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     5401 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     5461 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     5521 gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     5581 ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa
     5641 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     5701 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     5761 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     5821 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     5881 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     5941 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     6001 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     6061 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     6121 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     6181 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     6241 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     6301 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     6361 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     6421 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     6481 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     6541 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     6601 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     6661 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     6721 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     6781 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     6841 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     6901 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     6961 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     7021 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     7081 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     7141 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     7201 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     7261 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     7321 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     7381 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     7441 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     7501 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     7561 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     7621 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     7681 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     7741 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     7801 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     7861 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     7921 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     7981 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     8041 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     8101 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     8161 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     8221 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     8281 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     8341 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     8401 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     8461 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     8521 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     8581 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     8641 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     8701 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     8761 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     8821 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     8881 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     8941 aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt
//
