LOCUS       V019060                 7959 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V019060
VERSION     V019060
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     promoter        50..1228
                     /note="strong constitutive promoter for human elongation
                     factor EF-1α"
                     /label="EF-1α promoter"
     CDS             1281..2630
                     /label="BECN1(NM_003766)"
                     /note="BECN1(NM_003766)"
                     /gene="BECN1"
     CDS             2646..2711
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    2735..3323
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    3389..3437
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      3639..4067
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        4081..4410
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             4477..5271
                     /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    5447..5568
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(5617..5633)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    5641..5657
                     /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(5665..5695)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    5710..5731
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6019..6607)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(6778..7638)
                     /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(7639..7743)
                     /gene="bla"
                     /label="AmpR promoter"
ORIGIN
        1 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg gctccggtgc
       61 ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg
      121 caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta
      181 ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt
      241 gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc
      301 ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacctg
      361 gctgcagtac gtgattcttg atcccgagct tcgggttgga agtgggtggg agagttcgag
      421 gccttgcgct taaggagccc cttcgcctcg tgcttgagtt gaggcctggc ctgggcgctg
      481 gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt tcgataagtc
      541 tctagccatt taaaattttt gatgacctgc tgcgacgctt tttttctggc aagatagtct
      601 tgtaaatgcg ggccaagatc tgcacactgg tatttcggtt tttggggccg cgggcggcga
      661 cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag cgcggccacc
      721 gagaatcgga cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg gcctcgcgcc
      781 gccgtgtatc gccccgccct gggcggcaag gctggcccgg tcggcaccag ttgcgtgagc
      841 ggaaagatgg ccgcttcccg gccctgctgc agggagctca aaatggagga cgcggcgctc
      901 gggagagcgg gcgggtgagt cacccacaca aaggaaaagg gcctttccgt cctcagccgt
      961 cgcttcatgt gactccacgg agtaccgggc gccgtccagg cacctcgatt agttctcgag
     1021 cttttggagt acgtcgtctt taggttgggg ggaggggttt tatgcgatgg agtttcccca
     1081 cactgagtgg gtggagactg aagttaggcc agcttggcac ttgatgtaat tctccttgga
     1141 atttgccctt tttgagtttg gatcttggtt cattctcaag cctcagacag tggttcaaag
     1201 tttttttctt ccatttcagg tgtcgtgagg aattagcttg gtactggcta gcgtttaaac
     1261 ttaagcttgg taccgccacc atggaagggt ctaagacgtc caacaacagc accatgcagg
     1321 tgagcttcgt gtgccagcgc tgcagccagc ccctgaaact ggacacgagt ttcaagatcc
     1381 tggaccgtgt caccatccag gaactcacag ctccattact taccacagcc caggcgaaac
     1441 caggagagac ccaggaggaa gagactaact caggagagga gccatttatt gaaactcctc
     1501 gccaggatgg tgtctctcgc agattcatcc ccccagccag gatgatgtcc acagaaagtg
     1561 ccaacagctt cactctgatt ggggaggcat ctgatggcgg caccatggag aacctcagcc
     1621 gaagactgaa ggtcactggg gacctttttg acatcatgtc gggccagaca gatgtggatc
     1681 acccactctg tgaggaatgc acagatactc ttttagacca gctggacact cagctcaacg
     1741 tcactgaaaa tgagtgtcag aactacaaac gctgtttgga gatcttagag caaatgaatg
     1801 aggatgacag tgaacagtta cagatggagc taaaggagct ggcactagag gaggagaggc
     1861 tgatccagga gctggaagac gtggaaaaga accgcaagat agtggcagaa aatctcgaga
     1921 aggtccaggc tgaggctgag agactggatc aggaggaagc tcagtatcag agagaataca
     1981 gtgaatttaa acgacagcag ctggagctgg atgatgagct gaagagtgtt gaaaaccaga
     2041 tgcgttatgc ccagacgcag ctggataagc tgaagaaaac caacgtcttt aatgcaacct
     2101 tccacatctg gcacagtgga cagtttggca caatcaataa cttcaggctg ggtcgcctgc
     2161 ccagtgttcc cgtggaatgg aatgagatta atgctgcttg gggccagact gtgttgctgc
     2221 tccatgctct ggccaataag atgggtctga aatttcagag ataccgactt gttccttacg
     2281 gaaaccattc atatctggag tctctgacag acaaatctaa ggagctgccg ttatactgtt
     2341 ctggggggtt gcggtttttc tgggacaaca agtttgacca tgcaatggtg gctttcctgg
     2401 actgtgtgca gcagttcaaa gaagaggttg agaaaggcga gacacgtttt tgtcttccct
     2461 acaggatgga tgtggagaaa ggcaagattg aagacacagg aggcagtggc ggctcctatt
     2521 ccatcaaaac ccagtttaac tctgaggagc agtggacaaa agctctcaag ttcatgctga
     2581 cgaatcttaa gtggggtctt gcttgggtgt cctcacaatt ttataacaaa gtgcggccgc
     2641 tcgaggacta caaagaccat gacggtgatt ataaagatca tgacatcgac tacaaggatg
     2701 acgatgacaa gtagtgaggg cccgatatct cgacaatcaa cctctggatt acaaaatttg
     2761 tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc
     2821 tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta
     2881 taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt
     2941 ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca
     3001 gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc
     3061 ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt
     3121 gtcggggaag ctgacgtcct ttccatggct gctcgcctgt gttgccacct ggattctgcg
     3181 cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg
     3241 cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat
     3301 ctccctttgg gccgcctccc cgcctggaaa cgggggaggc taactgaaac acggaaggag
     3361 acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggg
     3421 tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac
     3481 cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc
     3541 cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat
     3601 agcagatctg cgcagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     3661 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     3721 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3781 agctctaaat cggggcatcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     3841 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     3901 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     3961 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgg ggatttcggc
     4021 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     4081 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     4141 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     4201 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     4261 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     4321 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     4381 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     4441 ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     4501 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     4561 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     4621 gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg
     4681 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     4741 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     4801 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     4861 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     4921 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     4981 gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat
     5041 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     5101 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     5161 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     5221 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     5281 tggggttcgc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     5341 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     5401 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     5461 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     5521 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     5581 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     5641 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     5701 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     5761 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     5821 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     5881 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     5941 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     6001 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     6061 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     6121 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     6181 ctttctccct tcgggaagcg tggcgctttc tcaatgctca cgctgtaggt atctcagttc
     6241 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     6301 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     6361 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     6421 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     6481 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     6541 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     6601 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     6661 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     6721 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     6781 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     6841 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     6901 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     6961 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     7021 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     7081 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     7141 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     7201 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     7261 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     7321 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     7381 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     7441 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     7501 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     7561 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     7621 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     7681 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     7741 gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc tcccgatccc
     7801 ctatggtcga ctctcagtac aatctgctct gatgccgcat agttaagcca gtatctgctc
     7861 cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc tacaacaagg
     7921 caaggcttga ccgacaattg catgaagaat ctgcttagg
//
