LOCUS       V040545                 7573 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V040545
VERSION     V040545
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     enhancer        235..614
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        615..818
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     promoter        863..881
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /label="T7 promoter"
     CDS             928..3009
                     /label="FZD8(NM_031866)"
                     /note="FZD8(NM_031866)"
                     /gene="FZD8"
     CDS             3016..3039
                     /codon_start=1
                     /product="FLAG® epitope tag, followed by an enterokinase
                     cleavage site"
                     /transl_table=1
                     /translation="DYKDDDDK"
                     /label="FLAG"
     CDS             3046..3135
                     /codon_start=1
                     /product="two Strep-Tag®II moieties connected by a linker
                     for enhanced binding to Strep-Tactin®,&nbsp;an engineered
                     form of streptavidin (Schmidt et al., 2013)"
                     /transl_table=1
                     /translation="SA,WSHPQFEK,GGGSGGGSGGSA,WSHPQFEK"
                     /label="Twin-Strep-tag"
     polyA_signal    3173..3397
                     /note="bovine growth hormone polyadenylation signal"
                     /label="bGH poly(A) signal"
     rep_origin      3443..3871
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        3885..4214
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             4281..5075
                     /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    5249..5370
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(5419..5435)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    5443..5459
                     /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(5467..5497)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    5512..5533
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(5821..6406)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(6577..7437)
                     /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(7438..7542)
                     /gene="bla"
                     /label="AmpR promoter"
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagctgaatt cgccaccatg gagtggggtt acctgttgga agtgacctcg
      961 ctgctggccg ccttggcgct gctgcagcgc tctagcggcg ctgcggccgc ctcggccaag
     1021 gagctggcat gccaagagat caccgtgccg ctgtgtaagg gcatcggcta caactacacc
     1081 tacatgccca atcagttcaa ccacgacacg caagacgagg cgggcctgga ggtgcaccag
     1141 ttctggccgc tggtggagat ccagtgctcg cccgatctca agttcttcct gtgcagcatg
     1201 tacacgccca tctgcctaga ggactacaag aagccgctgc cgccctgccg ctcggtgtgc
     1261 gagcgcgcca aggccggctg cgcgccgctc atgcgccagt acggcttcgc ctggcccgac
     1321 cgcatgcgct gcgaccggct gcccgagcaa ggcaaccctg acacgctgtg catggactac
     1381 aaccgcaccg acctaaccac cgccgcgccc agcccgccgc gccgcctgcc gccgccgccg
     1441 cccggcgagc agccgccttc gggcagcggc cacggccgcc cgccgggggc caggcccccg
     1501 caccgcggcg gcggcagggg cggtggcggc ggggacgcgg cggcgccccc agctcgcggc
     1561 ggcggcggtg gcgggaaggc gcggccccct ggcggcggcg cggctccctg cgagcccggg
     1621 tgccagtgcc gcgcgcctat ggtgagcgtg tccagcgagc gccacccgct ctacaaccgc
     1681 gtcaagacag gccagatcgc taactgcgcg ctgccctgcc acaacccctt tttcagccag
     1741 gacgagcgcg ccttcaccgt cttctggatc ggcctgtggt cggtgctctg cttcgtgtcc
     1801 accttcgcca ccgtctccac cttccttatc gacatggagc gcttcaagta cccggagcgg
     1861 cccattatct tcctctcggc ctgctacctc ttcgtgtcgg tgggctacct agtgcgcctg
     1921 gtggcgggcc acgagaaggt ggcgtgcagc ggtggcgcgc cgggcgcggg gggcgctggg
     1981 ggcgcgggcg gcgcggcggc gggcgcgggc gcggcgggcg cgggcgcggg cggcccgggc
     2041 gggcgcggcg agtacgagga gctgggcgcg gtggagcagc acgtgcgcta cgagaccacc
     2101 ggccccgcgc tgtgcaccgt ggtcttcttg ctggtctact tcttcggcat ggccagctcc
     2161 atctggtggg tgatcttgtc gctcacatgg ttcctggcgg ccggtatgaa gtggggcaac
     2221 gaagccatcg ccggctactc gcagtacttc cacctggccg cgtggcttgt gcccagcgtc
     2281 aagtccatcg cggtgctggc gctcagctcg gtggacggcg acccggtggc gggcatctgc
     2341 tacgtgggca accagagcct ggacaacctg cgcggcttcg tgctggcgcc gctggtcatc
     2401 tacctcttca tcggcaccat gttcctgctg gccggcttcg tgtccctctt ccgcatccgc
     2461 tcggtcatca agcaacagga cggccccacc aagacgcaca agctggagaa gctgatgatc
     2521 cgcctgggcc tgttcaccgt gctctacacc gtgcccgccg cggtggtggt cgcctgcctc
     2581 ttctacgagc agcacaaccg cccgcgctgg gaggccacgc acaactgccc gtgcctgcgg
     2641 gacctgcagc ccgaccaggc acgcaggccc gactacgccg tcttcatgct caagtacttc
     2701 atgtgcctag tggtgggcat cacctcgggc gtgtgggtct ggtccggcaa gacgctggag
     2761 tcctggcgct ccctgtgcac ccgctgctgc tgggccagca agggcgccgc ggtgggcggg
     2821 ggcgcgggcg ccacggccgc ggggggtggc ggcgggccgg ggggcggcgg cggcggggga
     2881 cccggcggcg gcggggggcc gggcggcggc gggggctccc tctacagcga cgtcagcact
     2941 ggcctgacgt ggcggtcggg cacggcgagc tccgtgtctt atccaaagca gatgccattg
     3001 tcccaggtcc tcgaggatta caaggatgac gacgataagg gatccagcgc ctggtcccac
     3061 cctcagtttg agaagggcgg aggctctggc ggcggaagcg gaggatctgc ttggagccac
     3121 ccccagttcg aaaagtgatg agggcccgtt taaacccgct gatcagcctc gactgtgcct
     3181 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     3241 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
     3301 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
     3361 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc
     3421 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     3481 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     3541 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     3601 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
     3661 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     3721 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     3781 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     3841 gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt
     3901 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     3961 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     4021 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     4081 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     4141 ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     4201 taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct gatcaagaga
     4261 caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg
     4321 cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg
     4381 ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt
     4441 ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg gccacgacgg
     4501 gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat
     4561 tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat
     4621 ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg
     4681 accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg
     4741 atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc
     4801 tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc
     4861 cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg
     4921 tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg
     4981 gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca
     5041 tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac
     5101 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga
     5161 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     5221 tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa
     5281 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     5341 tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta
     5401 gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     5461 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     5521 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     5581 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     5641 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     5701 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     5761 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     5821 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     5881 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     5941 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     6001 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     6061 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     6121 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     6181 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     6241 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     6301 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggttt
     6361 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     6421 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     6481 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     6541 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     6601 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     6661 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     6721 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     6781 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     6841 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     6901 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     6961 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     7021 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     7081 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     7141 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     7201 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     7261 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     7321 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     7381 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     7441 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     7501 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     7561 gccacctgac gtc
//
