LOCUS       V034682                 7564 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V034682
VERSION     V034682
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"
     regulatory      913..922
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             922..987
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             991..3084
                     /label="Atg7(NM_001012097)"
                     /note="Atg7(NM_001012097)"
                     /gene="Atg7"
     polyA_signal    3164..3388
                     /note="bovine growth hormone polyadenylation signal"
                     /label="bGH poly(A) signal"
     rep_origin      3434..3862
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        3876..4205
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             4272..5066
                     /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    5240..5361
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(5410..5426)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    5434..5450
                     /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(5458..5488)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    5503..5524
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(5812..6397)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(6568..7428)
                     /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(7429..7533)
                     /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 aagccaccat ggactacaaa gaccatgacg gtgattataa agatcatgac
      961 atcgactaca aggatgacga tgacaagctt atgggggacc ctggactgtc caagctgcag
     1021 tttgccccct ttaatagtgc cctggacgtt ggcttctggc acgaactgac ccagaagaag
     1081 ttgaacgagt atcgcctgga cgaggcaccc aaagacatca agggctatta ctacaatggt
     1141 gactctgccg gcctccccac ccgcttgaca ttggagttca gtgcttttga catgagcgcc
     1201 cccacgcctg cccgctgctg cccggccatg ggaaccctgc acaacaccaa cacacttgag
     1261 gcttttaaga ctgcagacaa gaagctcctt ctggagcagt cagccaatga gatctgggaa
     1321 gccataaagt caggtgctgc tctcgaaaac cccatgctcc tcaacaagtt tctgctcttg
     1381 accttcgcgg acctaaagaa gtaccacttc tactactggt tttgctgccc tgccctctgt
     1441 cttcctgaga gcatccctct aatccgggga cctgtgggct tggatcaaag gctttcacca
     1501 aaacagatcc aggccctgga gcatgcctac gatgacctgt gtcgaactga aggcgtcaca
     1561 gccctgccat acttcttatt caagtacgat gacgacactg tgctggtctc cttgctcaaa
     1621 cactacagtg atttcttcca aggtcaaagg acaaagttaa cagtcggtgt gtacgatccc
     1681 tgtaacctaa cccagcaccc tggatggcct ttgaggaatt ttttggtcct ggcagcccac
     1741 agatggagcg gcagtttcca gtctgttgaa gttctctgct ttcgggaccg caccatgcag
     1801 ggagcaagag atgtgacaca cagcatcatc tttgaagtga aacttccaga aatggcgttt
     1861 agcccagatt gtcctaaggc agttggctgg gagaagaacc agaaaggagg catgggtccg
     1921 aggatggtga acctcagcgg atgtatggac cccaaaaggc tggctgagtc atccgtggat
     1981 ctgaatctca agctgatgtg ctggcggttg gtccccacct tggacttgga caaggtcgtg
     2041 tctgtcaagt gcctgctgct gggagctggt accttggggt gtaatgtggc tagaacactg
     2101 atgggctggg gcgtcagaca cgtcacgttt gtggacaacg ccaagatctc ctactccaat
     2161 cccgtgaggc agcctctgta tgaatttgaa gattgtctag ggggtggcaa gcccaaggcc
     2221 ctggctgcag cagagcggct ccagaaaata tttcccggag tgaatgccag cgggttcaac
     2281 atgagcatcc ccatgcccgg acaccctgtg aacttctctg acgtcacgat ggagcaggcc
     2341 cgcagagatg tggagcagct ggaggaactc atcgatagcc acgatgtcat cttcctgcta
     2401 atggacacca gggagagccg atggcttcct actgttattg cagccagcaa gcgaaagctg
     2461 gtcatcaatg ctgccttggg gtttgacacc tttgttgtca tgagacatgg cctgaagaaa
     2521 cccaagcagc agggagccgg ggacctgtgc ccaagccatc ttgtagcacc tgctgacctg
     2581 ggctcctcac tttttgccaa catccctgga tacaagcttg gctgctactt ctgcaatgat
     2641 gtggtggctc caggagattc caccagagac cggactctgg accagcagtg cacagtgagc
     2701 cgcccagggc tggccgtgat tgcaggagcc ctggctgtgg agctgatggt ttctgtcttg
     2761 cagcatcctg aggggggcta cgccatcgcc agcagcagtg atgaccgcat gaatgagcct
     2821 cccacctcgc tggggcttgt acctcaccag atccggggct ttctgtcacg gttcgataat
     2881 gtccttcctg tcagcctggc atttgataaa tgtacagcct gttcatccaa agttcttgat
     2941 caatatgagc aagaaggatt caccttcctg gccaaggtgt ttaactcttc acattccttc
     3001 ttagaagatt tgactggtct tacactgctc caccaagaga cccaagctgc tgagatctgg
     3061 gacatgagtg acgaggagac cgtctgagaa ttctgcagat atccagcaca gtggcggccg
     3121 ctcgagtcta gagggcccgt ttaaacccac tgatcagcct cgactgtgcc ttctagttgc
     3181 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     3241 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     3301 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     3361 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     3421 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     3481 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     3541 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     3601 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     3661 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     3721 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     3781 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     3841 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt
     3901 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     3961 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     4021 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     4081 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     4141 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     4201 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag
     4261 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     4321 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     4381 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     4441 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     4501 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     4561 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     4621 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     4681 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     4741 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc
     4801 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     4861 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     4921 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     4981 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     5041 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     5101 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     5161 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     5221 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     5281 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     5341 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     5401 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     5461 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     5521 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     5581 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     5641 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     5701 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     5761 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     5821 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     5881 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     5941 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     6001 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     6061 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     6121 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     6181 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     6241 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     6301 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtt tttttgtttg
     6361 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     6421 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     6481 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     6541 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     6601 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     6661 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     6721 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     6781 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     6841 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     6901 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     6961 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     7021 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     7081 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     7141 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     7201 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     7261 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     7321 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     7381 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     7441 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     7501 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     7561 cgtc
//
