LOCUS       V025739                 8196 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V025739
VERSION     V025739
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             756..2876
                     /label="Appl1(NM_001399084)"
                     /note="Appl1(NM_001399084)"
                     /gene="Appl1"
     CDS             2883..2948
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    2972..3560
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    3626..3674
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      3876..4304
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        4318..4647
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             4714..5508
                     /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    5684..5805
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(5854..5870)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    5878..5894
                     /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(5902..5932)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    5947..5968
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6256..6844)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7015..7875)
                     /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(7876..7980)
                     /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 ctcgaattcg ccaccatgcc ggggatcgac aagctgccca
      781 ttgaagagac cctggaggac agtccgcaga caaggtcttt actgggcgta tttgaagaag
      841 atgctactgc tatttctaat tatatgaacc aattgtatca agctatgcat cggatttatg
      901 atgcacagaa tgaattaagt gcagcaactc acctgacttc gaaacttttg aaagaatatg
      961 agaagcagcg ttttcctctg gggggtgatg atgaagttat gagctctact ttgcagcagt
     1021 tttccaaggt catagatgag cttagttctt gtcatgcagt tctttcaact caactcgctg
     1081 atgccatgat gttccccatt tcccagttta aagaaagaga tctgaaagaa atattgacac
     1141 taaaagaggt atttcagatt gctagtaatg atcatgatgc cgcaattaat aggtacagcc
     1201 ggttgtcaaa aaagagagaa aatgacaagg taaagtatga agtaacagag gatgtgtata
     1261 cttccaggaa aaagcaacac cagaccatga tgcattattt ttgtgcacta aatactcttc
     1321 agtacaagaa gaaaatagct ttgttagaac ctctgcttgg atacatgcag gctcagataa
     1381 gtttctttaa gatgggttct gaaaatctca atggacaatt ggaagaattt ttagctaaca
     1441 ttggaacaag tgtacagaat gtgcgaaggg aaatggatgg ggatgtggag accatgcaac
     1501 agacaataga agacttggaa gtagccagtg accctttata tctgcctgac ccagatccca
     1561 ccaagtttcc cgtcaatcga aatttaaccc gaaaggctgg atacctaaat gctaggaata
     1621 aaacaggctt ggtgtcatct acctgggata gacagtttta cttcacgcag ggtggaaact
     1681 taatgagtca agcccgtgga gatgtggcag gaggcctagc catggacata gacaactgtt
     1741 cagtgatggc tgtggactgc gaagacagac gatactgttt tcagatcact tcttttgatg
     1801 gaaaaaaatc ttcaattttg caagcagaga gtaaaaaaga ccatgaagag tggatctgta
     1861 caataaacaa catatctaaa caaatatatt tgagtgaaaa tccagaggaa accgctgcac
     1921 gagtgaatca gtctgctcta gaagctgtca ctccttcccc atctttccag cagagacacg
     1981 agagcctgcg tccaggagga caatctcgac caccaacagc acgaaccagc agttcaggat
     2041 ccttgggatc agagtctaca aatttggctg ccctctctct agattctctt gttgctccag
     2101 atacaccaat acagtttgat ataatttctc ctgtgtgtga agatcagcct ggccaggcaa
     2161 aagcctttgg tcaaggaggc aggcgtacaa atccatttgg agaatctgga ggaagcacaa
     2221 aatcagaaac tgaagactct attcttcatc agttgtttat tgtccgattc cttggttcta
     2281 tggaggtaaa gtcagatgac catccagatg ttgtctatga aacaatgcgt cagatcttag
     2341 ctgctcgggc catccacaac atcttccgta tgacagaatc acatttatta gtcacttgtg
     2401 actgtctgaa gttaattgat ccacagaccc aagttacaag gctcacattt ccattaccat
     2461 atgtagttct gtatgctacg caccaggaaa ataaacggct ttttggattt gttctccgga
     2521 cctcaggcgg aagaagtgag agtaatctgt catcagtctg ctatatattt gagtccaaca
     2581 atgaggggga aaagatttgt gattctgttg gattggcaaa acagatagct ttgcatgctg
     2641 aactggatcg tagggcatca gaaaaacaaa aggaaataga gagagtaaaa gagaaacaac
     2701 agaaagaact cagcaaacag aaagagatcg agaaggactt ggaagaacag agtcggctga
     2761 tagctgcctc cagtagacca agccagtctg gtagcgaggg ccagcttgtt cttagcagta
     2821 gccaatcaga ggagagtgat ttgggagaag aaggaaaaaa aagagagtcg gaagcgggat
     2881 ccgactacaa agaccatgac ggtgattata aagatcatga catcgactac aaggatgacg
     2941 atgacaagta gtgagggccc gatatctcga caatcaacct ctggattaca aaatttgtga
     3001 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt
     3061 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa
     3121 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt
     3181 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct
     3241 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg
     3301 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc
     3361 ggggaagctg acgtcctttc catggctgct cgcctgtgtt gccacctgga ttctgcgcgg
     3421 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct
     3481 gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc
     3541 cctttgggcc gcctccccgc ctggaaacgg gggaggctaa ctgaaacacg gaaggagaca
     3601 ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacgggtgt
     3661 tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc
     3721 accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc
     3781 aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc
     3841 agatctgcgc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
     3901 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     3961 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     4021 tctaaatcgg ggcatccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     4081 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     4141 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     4201 actcaaccct atctcggtct attcttttga tttataaggg attttgggga tttcggccta
     4261 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
     4321 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     4381 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     4441 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     4501 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     4561 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc
     4621 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga
     4681 tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     4741 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     4801 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     4861 aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg
     4921 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     4981 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     5041 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     5101 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     5161 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     5221 ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc
     5281 gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt
     5341 ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct
     5401 gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc
     5461 gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg
     5521 ggttcgcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     5581 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     5641 cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc
     5701 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     5761 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc
     5821 gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     5881 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     5941 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     6001 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     6061 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     6121 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     6181 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     6241 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     6301 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     6361 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     6421 tctcccttcg ggaagcgtgg cgctttctca atgctcacgc tgtaggtatc tcagttcggt
     6481 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     6541 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     6601 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     6661 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
     6721 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     6781 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     6841 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     6901 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     6961 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     7021 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     7081 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     7141 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     7201 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     7261 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     7321 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     7381 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     7441 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     7501 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     7561 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     7621 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     7681 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     7741 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     7801 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     7861 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     7921 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     7981 cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta
     8041 tggtcgactc tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct
     8101 gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa
     8161 ggcttgaccg acaattgcat gaagaatctg cttagg
//
