LOCUS V040559 7432 bp DNA circular SYN 01-JAN-1980 DEFINITION synthetic circular DNA. ACCESSION V040559 VERSION V040559 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..2868 /label="FZD1(NM_003505)" /note="FZD1(NM_003505)" /gene="FZD1" CDS 2875..2898 /codon_start=1 /product="FLAG® epitope tag, followed by an enterokinase cleavage site" /transl_table=1 /translation="DYKDDDDK" /label="FLAG" CDS 2905..2994 /codon_start=1 /product="two Strep-Tag®II moieties connected by a linker for enhanced binding to Strep-Tactin®, an engineered form of streptavidin (Schmidt et al., 2013)" /transl_table=1 /translation="SA,WSHPQFEK,GGGSGGGSGGSA,WSHPQFEK" /label="Twin-Strep-tag" polyA_signal 3032..3256 /note="bovine growth hormone polyadenylation signal" /label="bGH poly(A) signal" rep_origin 3302..3730 /direction=RIGHT /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" /label="f1 ori" promoter 3744..4073 /note="SV40 enhancer and early promoter" /label="SV40 promoter" CDS 4140..4934 /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 5108..5229 /note="SV40 polyadenylation signal" /label="SV40 poly(A) signal" primer_bind complement(5278..5294) /note="common sequencing primer, one of multiple similar variants" /label="M13 rev" protein_bind 5302..5318 /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(5326..5356) /note="promoter for the E. coli lac operon" /label="lac promoter" protein_bind 5371..5392 /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." /label="CAP binding site" rep_origin complement(5680..6265) /direction=LEFT /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" /label="ori" CDS complement(6436..7296) /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(7297..7401) /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 gctgaggagg aggcgcctaa gaagtcccgg 961 gccgccggcg gtggcgcgag ctgggaactt tgtgccgggg cgctctcggc ccggctggcg 1021 gaggagggca gcggggacgc cggtggccgc cgccgcccgc cagttgaccc ccggcgattg 1081 gcgcgccagc tgctgctgct gctttggctg ctggaggctc cgctgctgct gggggtccgg 1141 gcccaggcgg cgggccaggg gccaggccag gggcccgggc cggggcagca accgccgccg 1201 ccgcctcagc agcaacagag cgggcagcag tacaacggcg agcggggcat ctccgtcccg 1261 gaccacggct attgccagcc catctccatc ccgctgtgca cggacatcgc gtacaaccag 1321 accatcatgc ccaacctgct gggccacacg aaccaggagg acgcgggcct ggaggtgcac 1381 cagttctacc ctctagtgaa agtgcagtgt tccgctgagc tcaagttctt cctgtgctcc 1441 atgtacgcgc ccgtgtgcac cgtgctagag caggcgctgc cgccctgccg ctccctgtgc 1501 gagcgcgcgc gccagggctg cgaggcgctc atgaacaagt tcggcttcca gtggccagac 1561 acgctcaagt gtgagaagtt cccggtgcac ggcgccggcg agctgtgcgt gggccagaac 1621 acgtccgaca agggcacccc gacgccctcg ctgcttccag agttctggac cagcaaccct 1681 cagcacggcg gcggagggca ccgtggcggc ttcccggggg gcgccggcgc gtcggagcga 1741 ggcaagttct cctgcccgcg cgccctcaag gtgccctcct acctcaacta ccacttcctg 1801 ggggagaagg actgcggcgc accttgtgag ccgaccaagg tgtatgggct catgtacttc 1861 gggcccgagg agctgcgctt ctcgcgcacc tggattggca tttggtcagt gctgtgctgc 1921 gcctccacgc tcttcacggt gcttacgtac ctggtggaca tgcggcgctt cagctacccg 1981 gagcggccca tcatcttctt gtccggctgt tacacggccg tggccgtggc ctacatcgcc 2041 ggcttcctcc tggaagaccg agtggtgtgt aatgacaagt tcgccgagga cggggcacgc 2101 actgtggcgc agggcaccaa gaaggagggc tgcaccatcc tcttcatgat gctctacttc 2161 ttcagcatgg ccagctccat ctggtgggtg atcctgtcgc tcacctggtt cctggcggct 2221 ggcatgaagt ggggccacga ggccatcgaa gccaactcac agtattttca cctggccgcc 2281 tgggctgtgc cggccatcaa gaccatcacc atcctggcgc tgggccaggt ggacggcgat 2341 gtgctgagcg gagtgtgctt cgtggggctt aacaacgtgg acgcgctgcg tggcttcgtg 2401 ctggcgcccc tcttcgtgta cctgtttatc ggcacgtcct ttctgctggc cggctttgtg 2461 tcgctcttcc gcatccgcac catcatgaag cacgatggca ccaagaccga gaagctggag 2521 aagctcatgg tgcgcattgg cgtcttcagc gtgctgtaca ctgtgccagc caccatcgtc 2581 atcgcctgct acttctacga gcaggccttc cgggaccagt gggaacgcag ctgggtggcc 2641 cagagctgca agagctacgc tatcccctgc cctcacctcc aggcgggcgg aggcgccccg 2701 ccgcacccgc ccatgagccc ggacttcacg gtcttcatga ttaagtacct tatgacgctg 2761 atcgtgggca tcacgtcggg cttctggatc tggtccggca agaccctcaa ctcctggagg 2821 aagttctaca cgaggctcac caacagcaaa caaggggaga ctacagtcct cgaggattac 2881 aaggatgacg acgataaggg atccagcgcc tggtcccacc ctcagtttga gaagggcgga 2941 ggctctggcg gcggaagcgg aggatctgct tggagccacc cccagttcga aaagtgatga 3001 gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt 3061 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc 3121 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt 3181 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat 3241 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc 3301 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc 3361 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc 3421 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt 3481 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg 3541 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt 3601 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta 3661 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt 3721 aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc 3781 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt 3841 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca 3901 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc 3961 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg 4021 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc 4081 cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca 4141 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg 4201 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag 4261 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc 4321 aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc 4381 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg 4441 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc 4501 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca 4561 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag 4621 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg 4681 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg 4741 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca 4801 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc 4861 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg 4921 acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct 4981 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt 5041 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc 5101 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa 5161 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa 5221 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc 5281 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg 5341 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt 5401 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg 5461 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga 5521 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat 5581 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca 5641 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc 5701 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata 5761 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc 5821 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc 5881 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga 5941 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc 6001 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag 6061 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag 6121 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag 6181 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtttt tttgtttgca agcagcagat 6241 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc 6301 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt 6361 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta 6421 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct 6481 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg 6541 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga 6601 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt 6661 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt 6721 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt 6781 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat 6841 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc 6901 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc 6961 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat 7021 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag 7081 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt 7141 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc 7201 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa 7261 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg 7321 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa 7381 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc //