LOCUS       V047257                 9054 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V047257
VERSION     V047257
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             756..3734
                     /label="Ephb3(NM_010143)"
                     /note="Ephb3(NM_010143)"
                     /gene="Ephb3"
     CDS             3741..3806
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    3830..4418
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    4484..4532
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      4734..5162
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        5176..5505
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             5572..6366
                     /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    6542..6663
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(6712..6728)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6736..6752
                     /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(6760..6790)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6805..6826
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7114..7702)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7873..8733)
                     /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(8734..8838)
                     /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 ctcgaattcg ccaccatggc cggagcccgc ccgccgccgg
      781 ggcttctgcc gctgctcgct ccgctgctgc tgccgctgct gcttcccgcc ggctgctggg
      841 cgctggaaga gactctcatg gacacgaaat gggtgacgtc tgagctggca tggacatctc
      901 atccagagag tgggtgggaa gaagtaagcg gctatgatga agccatgaat cctatccgca
      961 cgtatcaggt gtgtaatgtg cgtgagtcga gccagaacaa ctggctgcgg acgggtttca
     1021 tctggcggcg ggaagtccag cgcgtctacg tggagctgaa gttcaccgta agagactgta
     1081 acagcattcc caacatccct ggctcctgca aggaaacctt caaccttttc tactacgagg
     1141 ctgatagcga tgtggcctca gcctcctctc ccttctggat ggagaacccc tacgtgaaag
     1201 tggacaccat tgcgccagat gagagcttct cgcggctcga tgccgggcgt gtcaacacca
     1261 aggtgcgcag cttcgggccg ctttccaagg ctggcttcta cttggccttc caggaccaag
     1321 gtgcctgcat gtcactcatc tctgtgcgtg ccttctacaa gaagtgtgca tccaccactg
     1381 caggcttcgc actcttcccc gagaccctca cgggggctga gcccacttcg ctggtcattg
     1441 cccccggcac ctgcatcgct aacgctgtgg aggtctctgt accactcaag ctctactgca
     1501 atggcgacgg ggagtggatg gtgcccgtcg gtgcctgcac ctgcgctact ggccatgagc
     1561 cagccgccaa ggagtcccag tgccgtgcct gtcctcctgg gagttacaaa gcaaagcaag
     1621 gagaagggcc ctgcctcccc tgtcccccca atagccgcac cacctcgccg gctgccagca
     1681 tctgcacctg ccataataac ttctaccgtg cagactcaga ctcggccgac agcgcctgca
     1741 ccacggtgcc atctccaccc cggggtgtga tctccaatgt gaatgagacc tcgctaatcc
     1801 tcgaatggag tgagccccgg gaccttggcg gacgagatga cctcctttat aatgttatct
     1861 gcaagaagtg ccgtgggagc tctggggctg ggggtccggc aacctgttca cgctgcgatg
     1921 acaacgtaga gtttgtacct cggcagctgg gcctgacaga gcgccgggtc cacattagcc
     1981 acctgttggc ccacacacgc tacacctttg aggtgcaggc tgtcaacggt gtctcaggca
     2041 aaagcccttt gccaccccgc tatgcagctg tgaatatcac caccaaccag gctgctccat
     2101 cagaagtgcc tacactccac ctgcacagca gctcggggag cagcctgacc ctgtcctggg
     2161 cacccccaga gcggcctaat ggagtcatct tggactatga gatgaagtac tttgagaaga
     2221 gtaaaggcat cgcctccact gtaaccagcc agaagaactc cgtacaactg gacgggctgc
     2281 agcctgatgc ccgctacgta gttcaggtcc gggctcgcac agtagcgggt tacggacagt
     2341 atagccaccc ggctgagttt gagaccacaa gtgaaagagg ttcaggtgcc cagcagcttc
     2401 aagaacagct tcccctgatt gtgggatcca ccgtagccgg gtttgtcttc atggtggtcg
     2461 tcgtggtcat cgctcttgtc tgcctcagga agcagcgcca tggccccgat gcagaataca
     2521 cagagaagct gcagcaatac attgctcctg ggatgaaagt ttacattgac ccctttacct
     2581 atgaggatcc caatgaggcc gtccgagagt tcgccaagga gatcgatgtg tcctgtgtca
     2641 agatcgagga ggtgattgga gctggggagt ttggggaagt gtgccggggt cgactgaaac
     2701 tgcccggccg ccgggaggtg ttcgtggcta tcaagacgct gaaggtggga tacacggaga
     2761 ggcagcggcg ggacttccta agtgaggctt ccattatggg tcagtttgac catcccaata
     2821 taatccgtct agagggtgtg gtcaccaaaa gtcgtccagt catgatcctc actgagttca
     2881 tggagaactg tgccctggac tcctttctac ggctcaatga cggacagttc acggtcatcc
     2941 agcttgtggg catgttacga ggcattgctg ctggcatgaa gtacttgtct gagatgaact
     3001 acgtgcaccg tgaccttgct gcccgaaaca tccttgtcaa cagcaacttg gtctgcaaag
     3061 tctctgactt tggcctctcc cgcttcctgg aggatgaccc atcagacccc acctacacca
     3121 gctccctggg tgggaagatc cctatccgct ggaccgcccc ggaggccata gcctatcgga
     3181 agtttacgtc tgccagcgat gtctggagct atgggattgt catgtgggag gtcatgagct
     3241 acggagagcg accctactgg gatatgagca accaggatgt cattaatgct gtagagcaag
     3301 actatcggtt accacccccc atggactgcc ccaccgcgct gcaccagctc atgctggact
     3361 gttgggtgcg ggaccggaac ctcaggccca agttctctca aatcgtcaac acactagaca
     3421 aacttatccg caatgctgcc agcctcaagg tcaccgccag tgccccatct ggcatgtccc
     3481 agcccctcct ggaccgcaca gtcccagatt atacgacctt cacgacagtg ggtgactggc
     3541 tagatgccat caagatgggg aggtataaag agagctttgt cggtgctggg tttgcatcct
     3601 ttgacctggt ggcccagatg actgcagaag atctgctaag gattggggtc actttggctg
     3661 ggcaccagaa gaagatcctc tgcagtatcc aggacatgcg gctgcagatg aaccagaccc
     3721 tgcctgtgca ggtcggatcc gactacaaag accatgacgg tgattataaa gatcatgaca
     3781 tcgactacaa ggatgacgat gacaagtagt gagggcccga tatctcgaca atcaacctct
     3841 ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct
     3901 atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat
     3961 tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt
     4021 caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat
     4081 tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc
     4141 ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga
     4201 caattccgtg gtgttgtcgg ggaagctgac gtcctttcca tggctgctcg cctgtgttgc
     4261 cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga
     4321 ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc
     4381 tcagacgagt cggatctccc tttgggccgc ctccccgcct ggaaacgggg gaggctaact
     4441 gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag
     4501 aataaaacgc acgggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg
     4561 gcactctgtc gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt
     4621 ttccccaccc caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc
     4681 ggcaggccct gccatagcag atctgcgcag ctggggctct agggggtatc cccacgcgcc
     4741 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     4801 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     4861 cggctttccc cgtcaagctc taaatcgggg catcccttta gggttccgat ttagtgcttt
     4921 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     4981 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     5041 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     5101 tttggggatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     5161 ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
     5221 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     5281 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     5341 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     5401 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc
     5461 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct
     5521 tgtatatcca ttttcggatc tgatcaagag acaggatgag gatcgtttcg catgattgaa
     5581 caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac
     5641 tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg
     5701 cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag
     5761 gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt
     5821 gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg
     5881 tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg
     5941 catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga
     6001 gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag
     6061 gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat
     6121 ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt
     6181 tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg
     6241 gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt
     6301 tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc
     6361 ttctgagcgg gactctgggg ttcgcgaaat gaccgaccaa gcgacgccca acctgccatc
     6421 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     6481 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     6541 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     6601 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     6661 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     6721 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     6781 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     6841 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     6901 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     6961 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     7021 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     7081 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     7141 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     7201 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     7261 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcaat gctcacgctg
     7321 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     7381 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     7441 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     7501 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     7561 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     7621 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     7681 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     7741 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     7801 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     7861 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     7921 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     7981 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     8041 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     8101 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     8161 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     8221 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     8281 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     8341 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     8401 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     8461 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     8521 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     8581 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     8641 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     8701 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     8761 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     8821 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg
     8881 agatctcccg atcccctatg gtcgactctc agtacaatct gctctgatgc cgcatagtta
     8941 agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt
     9001 taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct tagg
//
