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®,&nbsp;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
//
