LOCUS       V023636                 7838 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V023636
VERSION     V023636
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             779..2518
                     /label="MCOLN1(NM_020533)"
                     /note="MCOLN1(NM_020533)"
                     /gene="MCOLN1"
     CDS             2525..2590
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    2614..3202
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    3268..3316
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      3518..3946
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        3960..4289
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             4356..5150
                     /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    5326..5447
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(5496..5512)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    5520..5536
                     /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(5544..5574)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    5589..5610
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(5898..6486)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(6657..7517)
                     /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(7518..7622)
                     /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 ctcggatcca ctagtccagt gtggtggaat tcgccaccat
      781 gacagccccg gcgggtccgc gcggctcaga gaccgagcgg cttctgaccc ccaaccccgg
      841 gtatgggacc caggcggggc cttcaccggc ccctccgaca cccccagaag aggaagacct
      901 tcgccgtcgt ctcaaatact ttttcatgag tccctgcgac aagtttcgag ccaagggccg
      961 caagccctgc aagctgatgc tgcaagtggt caagatcctg gtggtcacgg tgcagctcat
     1021 cctgtttggg ctcagtaatc agctggctgt gacattccgg gaagagaaca ccatcgcctt
     1081 ccgacacctc ttcctgctgg gctactcgga cggagcggat gacaccttcg cagcctacac
     1141 gcgggagcag ctgtaccagg ccatcttcca tgctgtggac cagtacctgg cgttgcctga
     1201 cgtgtcactg ggccggtatg cgtatgtccg tggtgggggt gacccttgga ccaatggctc
     1261 agggcttgct ctctgccagc ggtactacca ccgaggccac gtggacccgg ccaacgacac
     1321 atttgacatt gatccgatgg tggttactga ctgcatccag gtggatcccc ccgagcggcc
     1381 ccctccgccc cccagcgacg atctcaccct cttggaaagc agctccagtt acaagaacct
     1441 cacgctcaaa ttccacaagc tggtcaatgt caccatccac ttccggctga agaccattaa
     1501 cctccagagc ctcatcaata atgagatccc ggactgctat accttcagcg tcctgatcac
     1561 gtttgacaac aaagcacaca gtgggcggat ccccatcagc ctggagaccc aggcccacat
     1621 ccaggagtgt aagcacccca gtgtcttcca gcacggagac aacagcttcc ggctcctgtt
     1681 tgacgtggtg gtcatcctca cctgctccct gtccttcctc ctctgcgccc gctcactcct
     1741 tcgaggcttc ctgctgcaga acgagtttgt ggggttcatg tggcggcagc ggggacgggt
     1801 catcagcctg tgggagcggc tggaatttgt caatggctgg tacatcctgc tcgtcaccag
     1861 cgatgtgctc accatctcgg gcaccatcat gaagatcggc atcgaggcca agaacttggc
     1921 gagctacgac gtctgcagca tcctcctggg cacctcgacg ctgctggtgt gggtgggcgt
     1981 gatccgctac ctgaccttct tccacaacta caatatcctc atcgccacac tgcgggtggc
     2041 cctgcccagc gtcatgcgct tctgctgctg cgtggctgtc atctacctgg gctactgctt
     2101 ctgtggctgg atcgtgctgg ggccctatca tgtgaagttc cgctcactct ccatggtgtc
     2161 tgagtgcctg ttctcgctca tcaatgggga cgacatgttt gtgacgttcg ccgccatgca
     2221 ggcgcagcag ggccgcagca gcctggtgtg gctcttctcc cagctctacc tttactcctt
     2281 catcagcctc ttcatctaca tggtgctcag cctcttcatc gcgctcatca ccggcgccta
     2341 cgacaccatc aagcatcccg gcggcgcagg cgcagaggag agcgagctgc aggcctacat
     2401 cgcacagtgc caggacagcc ccacctccgg caagttccgc cgcgggagcg gctcggcctg
     2461 cagccttctc tgctgctgcg gaagggaccc ctcggaggag cattcgctgc tggtgaatct
     2521 cgaggactac aaagaccatg acggtgatta taaagatcat gacatcgact acaaggatga
     2581 cgatgacaag tagtgagggc ccgatatctc gacaatcaac ctctggatta caaaatttgt
     2641 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     2701 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     2761 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     2821 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     2881 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     2941 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     3001 tcggggaagc tgacgtcctt tccatggctg ctcgcctgtg ttgccacctg gattctgcgc
     3061 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     3121 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     3181 tccctttggg ccgcctcccc gcctggaaac gggggaggct aactgaaaca cggaaggaga
     3241 caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa acgcacgggt
     3301 gttgggtcgt ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc
     3361 ccaccgagac cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc
     3421 ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata
     3481 gcagatctgc gcagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta
     3541 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     3601 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     3661 gctctaaatc ggggcatccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     3721 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     3781 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     3841 acactcaacc ctatctcggt ctattctttt gatttataag ggattttggg gatttcggcc
     3901 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg
     3961 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     4021 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
     4081 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
     4141 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     4201 ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag
     4261 gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg
     4321 gatctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     4381 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     4441 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     4501 tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt
     4561 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     4621 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     4681 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     4741 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     4801 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     4861 aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg
     4921 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     4981 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     5041 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     5101 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     5161 ggggttcgcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc
     5221 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg
     5281 atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca
     5341 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
     5401 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata
     5461 ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat
     5521 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg
     5581 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag
     5641 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     5701 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     5761 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     5821 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     5881 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     5941 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     6001 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     6061 tttctccctt cgggaagcgt ggcgctttct caatgctcac gctgtaggta tctcagttcg
     6121 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     6181 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     6241 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     6301 ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct
     6361 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     6421 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     6481 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     6541 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     6601 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     6661 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     6721 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     6781 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     6841 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     6901 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     6961 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     7021 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     7081 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     7141 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     7201 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     7261 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     7321 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     7381 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     7441 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     7501 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     7561 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     7621 cgcacatttc cccgaaaagt gccacctgac gtcgacggat cgggagatct cccgatcccc
     7681 tatggtcgac tctcagtaca atctgctctg atgccgcata gttaagccag tatctgctcc
     7741 ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa aatttaagct acaacaaggc
     7801 aaggcttgac cgacaattgc atgaagaatc tgcttagg
//
