LOCUS V040020 6442 bp DNA circular SYN 01-JAN-1980 DEFINITION synthetic circular DNA. ACCESSION V040020 VERSION V040020 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..1878 /label="MC1R(NM_002386)" /note="MC1R(NM_002386)" /gene="MC1R" CDS 1885..1908 /codon_start=1 /product="FLAG® epitope tag, followed by an enterokinase cleavage site" /transl_table=1 /translation="DYKDDDDK" /label="FLAG" CDS 1915..2004 /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 2042..2266 /note="bovine growth hormone polyadenylation signal" /label="bGH poly(A) signal" rep_origin 2312..2740 /direction=RIGHT /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" /label="f1 ori" promoter 2754..3083 /note="SV40 enhancer and early promoter" /label="SV40 promoter" CDS 3150..3944 /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 4118..4239 /note="SV40 polyadenylation signal" /label="SV40 poly(A) signal" primer_bind complement(4288..4304) /note="common sequencing primer, one of multiple similar variants" /label="M13 rev" protein_bind 4312..4328 /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(4336..4366) /note="promoter for the E. coli lac operon" /label="lac promoter" protein_bind 4381..4402 /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." /label="CAP binding site" rep_origin complement(4690..5275) /direction=LEFT /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" /label="ori" CDS complement(5446..6306) /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(6307..6411) /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 gctgtgcagg gatcccagag aagacttctg 961 ggctccctca actccacccc cacagccatc ccccagctgg ggctggctgc caaccagaca 1021 ggagcccggt gcctggaggt gtccatctct gacgggctct tcctcagcct ggggctggtg 1081 agcttggtgg agaacgcgct ggtggtggcc accatcgcca agaaccggaa cctgcactca 1141 cccatgtact gcttcatctg ctgcctggcc ttgtcggacc tgctggtgag cgggagcaac 1201 gtgctggaga cggccgtcat cctcctgctg gaggccggtg cactggtggc ccgggctgcg 1261 gtgctgcagc agctggacaa tgtcattgac gtgatcacct gcagctccat gctgtccagc 1321 ctctgcttcc tgggcgccat cgccgtggac cgctacatct ccatcttcta cgcactgcgc 1381 taccacagca tcgtgaccct gccgcgggcg cggcgagccg ttgcggccat ctgggtggcc 1441 agtgtcgtct tcagcacgct cttcatcgcc tactacgacc acgtggccgt cctgctgtgc 1501 ctcgtggtct tcttcctggc tatgctggtg ctcatggccg tgctgtacgt ccacatgctg 1561 gcccgggcct gccagcacgc ccagggcatc gcccggctcc acaagaggca gcgcccggtc 1621 caccagggct ttggccttaa aggcgctgtc accctcacca tcctgctggg cattttcttc 1681 ctctgctggg gccccttctt cctgcatctc acactcatcg tcctctgccc cgagcacccc 1741 acgtgcggct gcatcttcaa gaacttcaac ctctttctcg ccctcatcat ctgcaatgcc 1801 atcatcgacc ccctcatcta cgccttccac agccaggagc tccgcaggac gctcaaggag 1861 gtgctgacat gctcctggct cgaggattac aaggatgacg acgataaggg atccagcgcc 1921 tggtcccacc ctcagtttga gaagggcgga ggctctggcg gcggaagcgg aggatctgct 1981 tggagccacc cccagttcga aaagtgatga gggcccgttt aaacccgctg atcagcctcg 2041 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc 2101 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt 2161 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat 2221 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa 2281 agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg 2341 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct 2401 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta 2461 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa 2521 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct 2581 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc 2641 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg 2701 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc 2761 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc 2821 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc 2881 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc 2941 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt 3001 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt 3061 ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg 3121 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt 3181 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct 3241 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga 3301 ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg 3361 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact 3421 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg 3481 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct 3541 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg 3601 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt 3661 tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg 3721 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc 3781 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag 3841 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt 3901 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt 3961 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc 4021 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca 4081 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa 4141 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca 4201 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac 4261 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc 4321 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta 4381 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa 4441 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat 4501 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg 4561 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc 4621 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt 4681 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag 4741 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc 4801 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc 4861 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt 4921 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt 4981 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc 5041 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa 5101 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa 5161 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg 5221 tagcggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga 5281 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat 5341 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag 5401 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat 5461 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc 5521 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat 5581 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag 5641 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg 5701 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc 5761 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca 5821 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg 5881 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc 5941 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta 6001 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc 6061 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg 6121 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc 6181 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc 6241 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat 6301 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag 6361 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc 6421 ccgaaaagtg ccacctgacg tc //