LOCUS       V032765                 9978 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V032765
VERSION     V032765
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..4658
                     /label="AMBRA1(NM_001267782)"
                     /note="AMBRA1(NM_001267782)"
                     /gene="AMBRA1"
     CDS             4665..4730
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    4754..5342
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    5408..5456
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      5658..6086
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        6100..6429
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             6496..7290
                     /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    7466..7587
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(7636..7652)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    7660..7676
                     /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(7684..7714)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    7729..7750
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(8038..8626)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8797..9657)
                     /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(9658..9762)
                     /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 ccaccatgaa ggttgtccca gaaaagaatg
      781 ctgtccggat actctggggg cgagaacggg gtgctcgggc catgggagct cagcggcttc
      841 tgcaggagct ggtagaagat aaaacccggt ggatgaaatg ggagggcaag agagtagaac
      901 tgccggatag tccacgctct accttcttat tggccttcag cccagacagg actctcttag
      961 cctccaccca tgtgaaccat aatatctata ttacggaggt gaagactggc aagtgtgttc
     1021 attccctgat tggacaccgc cgtactccat ggtgtgtcac ttttcatccc accatctcag
     1081 gccttattgc ttctggctgc ctagatgggg aggttaggat ttgggattta cacggtggca
     1141 gtgaaagctg gttcacagat agcaacaatg ccattgcctc cctggctttc caccctacgg
     1201 ctcagctcct gctgattgcc actgccaatg agatccactt ctgggactgg agtcgacggg
     1261 aaccctttgc tgtggtgaag acagctagtg agatggaacg ggtccgtctg gtgagatttg
     1321 atccacttgg acactactta ctcacagcaa ttgttaaccc ctctaatcaa cagggtgatg
     1381 acgaaccaga gatccccata gatggaacag aattatccca ctaccgtcag cgtgccctcc
     1441 tgcaatcaca gccagttcgc cggacgcctc tcctccacaa tttcctgcac atgctgtcct
     1501 cccgctcttc tggcatccag accgagccct tccatccccc ggagcaggcc tcgtcaacgc
     1561 agcaggacca gggcctcctg aaccggccgt ctgccttcag tacagtccag agcagcactg
     1621 ccggcaacac gctccgcaac ctcagtctgg gtcctacccg ccgctctttg ggagggcctc
     1681 tgtctagcca cccttctagg tatcaccgag aaatagctcc tgggttgaca ggatctgagt
     1741 ggacccggac agtactcagt ctgaactccc gctctgaggc ggaatccatg cccccgccca
     1801 gaaccagtgc ctcttcggtg agtttgctgt ctgtgctgag acagcaggaa ggtggctctc
     1861 aggcatctgt gtacacttca gccacagaag ggaggggttt tccggcatca gggttggcaa
     1921 ctgagtcaga tggagggaat ggctccagcc aaaacaactc gggcagcatt cgccatgagc
     1981 ttcagtgtga cctgagacgc ttctttctgg agtatgaccg gcttcaggag ctggatcaga
     2041 gcctgagtgg ggaagctccc cagacccaac aggcccagga aatgctcaac aataacattg
     2101 aatctgagag gccaggccct tcccaccagc ccaccccaca cagcagtgag aacaactcca
     2161 acctgtcccg tggccacctg aatcgctgtc gtgcttgcca caatctcctg accttcaaca
     2221 acgataccct gcgctgggaa agaaccacac ctaactactc ctctggcgag gctagttcct
     2281 cttggcaggt ccccagctcc tttgagagtg tgccatcaag tggcagccag ttgccacctc
     2341 tcgagcggac tgagggccaa acgcccagct ccagcaggct ggagttgagc agctctgcta
     2401 gtccgcagga ggagaggact gtgggggtgg cctttaacca ggagacaggc cactgggaaa
     2461 gaatttacac ccagtccagc agatctggaa ctgtgtcaca ggaggcctta catcaggata
     2521 tgcctgagga gagctctgag gaggattcac tcaggagacg gagtcttgct ctgtcaccta
     2581 ggctggagta cagcggcgct atcttggctc actgcaagct ccgcctccca ggttcatgcc
     2641 attctcctgc ctcagcttcc caagtagctg ggactacggg agcccaccac cacgcccggc
     2701 taatttttgc atttttagta gagatggagt ttcaccatgt tagccaggct ggtctcgaac
     2761 tcctgacctc aggtgatctg cccacctcgg cctcccaaag tgctgggatt acaggcgtga
     2821 gccaccgcgc ctggccgagg ctgctggaat cttccctcat ttcattatcc cgttatgatg
     2881 gagcaggatc cagagagcac ccaatttacc cagacccagc gagattatct cctgctgcat
     2941 actacgccca gaggatgatc cagtatctct cacggagaga cagtattcgc cagcgctcca
     3001 tgcgctacca acagaaccgt ctccgttctt ccacctcctc ctcttcctca gacaaccagg
     3061 gtccatcagt agagggaacc gacttggaat ttgaggactt tgaggacaat ggtgacagat
     3121 ccaggcaccg agctccacgc aatgcccgga tgtctgcacc ttcgcttgga cgctttgtcc
     3181 caaggcgttt cttgctgcct gagtacttgc cttatgctgg gatttttcat gaacgtggac
     3241 agcctggctt ggctactcac tcttctgtta acagggtcct ggcaggggca gtgatcggtg
     3301 atggacagtc tgctgtggcc agtaacattg ccaatactac ctaccggctc cagtggtggg
     3361 acttcactaa gtttgacctc cctgaaatca gtaatgcttc cgtgaatgtg ctggtgcaga
     3421 actgcaagat ctacaatgat gccagctgtg acatttctgc agatggccag ctcctggcag
     3481 ctttcatccc cagcagccag aggggctttc ctgatgaagg catcctggca gtgtactccc
     3541 tggcccccca taacctgggc gaaatgctct acaccaagcg atttggtccc aatgccattt
     3601 cggtgagcct gtccccaatg ggcagatatg taatggtggg cttggcctca cgaaggatcc
     3661 tgctgcaccc ctccacagag cacatggtgg cccaggtctt caggctgcaa caggcccatg
     3721 gtggagagac ctccatgagg agagttttca acgtccttta tcccatgcct gccgaccagc
     3781 ggagacatgt cagtatcaac tctgcccgtt ggctgcctga gccagggctt ggcttggcct
     3841 atggtactaa caaaggagac ctggtgatct gccgaccaga ggccttaaac tctggtgttg
     3901 agtactactg ggaccagctg aacgagacgg tcttcactgt ccattccaac agcaggagca
     3961 gcgagcggcc tggaaccagc agagccacat ggaggacaga cagagacatg gggctgatga
     4021 atgccattgg gcttcagccc cggaaccctg ccacctcagt gacatctcag ggcacccaga
     4081 ctctggccct tcagctgcag aatgccgaaa cacagactga gagggaggtg ccggagccag
     4141 ggacagccgc ctcaggtcct ggtgaaggtg agggttcaga gtatggtgcc agtggagaag
     4201 atgcgctcag caggatccag aggctgatgg cggagggcgg catgacagcc gtggtgcagc
     4261 gggagcagag caccaccatg gcctccatgg gcggcttcgg caacaacatc atcgtcagcc
     4321 accgcattca ccgcagctct cagacgggca ctgaacctgg tgccgcccac acctcctcac
     4381 cccagccctc cacctctcgg ggactgctcc cagaggccgg gcaactggca gagcgaggcc
     4441 taagcccccg gacagcttcc tgggaccagc ctggtacccc tgggcgggag ccaacccagc
     4501 caaccctgcc ctcttcctcc cctgtcccca ttcctgtttc ccttcccagc gctgagggac
     4561 caaccctcca ctgcgagttg accaataaca accaccttct ggatggtggc agcagcaggg
     4621 gggacgctgc aggccctagg ggagaaccac ggaacagggg atccgactac aaagaccatg
     4681 acggtgatta taaagatcat gacatcgact acaaggatga cgatgacaag tagtgagggc
     4741 ccgatatctc gacaatcaac ctctggatta caaaatttgt gaaagattga ctggtattct
     4801 taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc
     4861 tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct
     4921 ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga
     4981 cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc
     5041 tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac
     5101 aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaagc tgacgtcctt
     5161 tccatggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt
     5221 cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc
     5281 tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc
     5341 gcctggaaac gggggaggct aactgaaaca cggaaggaga caataccgga aggaacccgc
     5401 gctatgacgg caataaaaag acagaataaa acgcacgggt gttgggtcgt ttgttcataa
     5461 acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg
     5521 ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc
     5581 agggctcgca gccaacgtcg gggcggcagg ccctgccata gcagatctgc gcagctgggg
     5641 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     5701 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     5761 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc ggggcatccc
     5821 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     5881 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     5941 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     6001 ctattctttt gatttataag ggattttggg gatttcggcc tattggttaa aaaatgagct
     6061 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
     6121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     6181 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     6241 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     6301 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     6361 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     6421 ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca agagacagga
     6481 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     6541 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     6601 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     6661 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     6721 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     6781 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     6841 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     6901 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     6961 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     7021 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     7081 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     7141 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     7201 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     7261 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgcg aaatgaccga
     7321 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     7381 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     7441 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     7501 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     7561 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     7621 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     7681 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     7741 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     7801 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     7861 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     7921 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     7981 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     8041 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     8101 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     8161 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     8221 ggcgctttct caatgctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     8281 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     8341 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     8401 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     8461 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     8521 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     8581 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     8641 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     8701 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     8761 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     8821 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     8881 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     8941 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     9001 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     9061 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     9121 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     9181 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     9241 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     9301 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     9361 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     9421 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     9481 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     9541 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     9601 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     9661 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     9721 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     9781 gccacctgac gtcgacggat cgggagatct cccgatcccc tatggtcgac tctcagtaca
     9841 atctgctctg atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc
     9901 gctgagtagt gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc
     9961 atgaagaatc tgcttagg
//
