LOCUS       V035612                 8655 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V035612
VERSION     V035612
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..3335
                     /label="Tgfbrap1(NM_001013025)"
                     /note="Tgfbrap1(NM_001013025)"
                     /gene="Tgfbrap1"
     CDS             3342..3407
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    3431..4019
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    4085..4133
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      4335..4763
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        4777..5106
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             5173..5967
                     /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    6143..6264
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(6313..6329)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6337..6353
                     /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(6361..6391)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6406..6427
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6715..7303)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7474..8334)
                     /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(8335..8439)
                     /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 ccaccatgat gaacatcaag gccttcacct
      781 tggtctctgc agtggaacgg gaactgctga tgggcgacag agaccacatc agcattgagt
      841 gtgtggagtg ctgtggcagg aatctttatg tcggcaccaa tgactgcttc atttaccatt
      901 tcctgctgga ggagaaggcc atgcctacag ggacagctac attcgttgcc accaaacagc
      961 tacacagaca cctgggcttc aagaaacctg tgaacgagct gtgtgcggcc tcagctctca
     1021 ataggctgct ggtgctctgt gacaactcta taaccctggt taacatgctg aacctggagc
     1081 cagtcccctc aggagcccgc atcaaagggg ctacaacatt tgcagtgaac gagagccccg
     1141 tgaatgggga cccgttctgc gtggaggtgt gcatcatttc cgtgaagcgc aggactgtcc
     1201 aaatgtttct ggtgtatgag gaccgagtgc agattgtcaa ggaagtgtcc accccagagc
     1261 agcctcttgc tgtggctgtg gatggctatt tcttgtgcct ggccttgacc acacaataca
     1321 tcatcctgaa ctacagcaca ggcctttccc aggacctgtt tccctactgc agtgaggaga
     1381 agccacccat cgtcaagagg ataggcagac aggagttcct gctggctggc ccaggaggct
     1441 tgggcatgtt tgccaccgtg gccgggattt cccagcgtgc ccctgtgcac tggtcagaaa
     1501 atgtgattgg ggcagctgtt tgcttcccct atgtcatagc gctcgatgac gaattcatca
     1561 cagttcacag catgttggat cagcagcaga aacagaccct gccctttaag gaggggcaca
     1621 tcctgcagga ctttgaagga agagtgattg ttgccacaag taaaggagtt tacatcttgg
     1681 ttccattacc tctggaaaaa caaatacagg atctgttagc aaaccggaga gtggaggagg
     1741 ctttggtctt ggcaaaagga gcccggagga acattccaaa ggagaaattt caggtaatgt
     1801 acaggaggat tctgcagcag gcaggcttta tacagtttgc acaactccag ttcctggaag
     1861 ccaaagagct cttcagaagc agccagctgg atgtccggga gctgatctct ctctacccct
     1921 ttctgctgcc cacctcctcc tccttcaccc gctctcaccc tcctcttcat gagtacgcag
     1981 acctgaacca gctgacccaa ggtgaccagg agaagatggc caagtgcaag cgcttcctca
     2041 tgagctacct gaatgaaatc cgcagcacag aggtagctaa tggctacaag gaagacatcg
     2101 acacggcctt gctcaagctg tatgcagagg ctgaccacga cagcctcctg gacctcttgg
     2161 tcactgagaa tttctgcttg ctgacagaca gtgctgcctg gctggagaag cacaaaaagt
     2221 actttgcact ggggctgctc tatcattaca acaaacaaga tgcctctgcg gttcagttgt
     2281 gggtgaacat tgtgaatggg gacatccagg actctacacg ctcggacctg tatgaataca
     2341 tcgttgattt ccttacctac tgcctagacc aagagctggt gtggacccat gctgactggc
     2401 tgctgcagaa aagtgaggag attggagttc aaattttcac caagagacct ttggacgagc
     2461 agcagcagac tagttttaat ccagacaaca ttattagttc ccttaagaaa taccctaaag
     2521 ccctggttaa gtatctggaa catctcgtga tcgacaggag attgcagaaa gaagagtacc
     2581 acacccacct tgccatcctc tacctggagg aagtgctgcg gcagagggta tccactgggg
     2641 gcaaagatgt agaagccact gagacacagg ccaaactccg gagactactc cagaagtctg
     2701 acttgtaccg cgtccacttg ttgaaagaga aagtccaggg tgccggcctg cccatggaaa
     2761 gtgccatcct ccacgggaag ctgggcgagc acgagaaggc cctgcacatc ctggtgcacg
     2821 agatggggga cttctctgca gctgaggact actgcctgtg gagctccgag ggtcagggtg
     2881 cagcctgccg tcagcgcctc ttccacacgt tgctcgccat gtacctccgt gcaggcccct
     2941 ctgcccagga tctgacagtg gccgctgtgg acttgctgaa ccatcatgcc agggaatttg
     3001 atgtcaccca ggtcctacag ctgctgccag acacctggtc cgtgcagctt ctctgcccat
     3061 tcctcatggg ggctatgagg gacagtatcc atgccaggag gaccacacag gtggcccttg
     3121 gcctggccaa atcagaaaac ctgatttaca tgtatgataa gatgaagctg aagggcaatg
     3181 cagtccggct ctcagaacga gagctttgcc agctctgcca gaaccctttc ggtgaacctg
     3241 tgtttgtcag atacccaaat ggagggcttg tgcacaccca ctgtgccgcc agcagacaca
     3301 cggcccccag ctcgcccagc cctggcactc ggactggatc cgactacaaa gaccatgacg
     3361 gtgattataa agatcatgac atcgactaca aggatgacga tgacaagtag tgagggcccg
     3421 atatctcgac aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa
     3481 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat
     3541 tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta
     3601 tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc
     3661 aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt
     3721 ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg
     3781 ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaagctga cgtcctttcc
     3841 atggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc
     3901 ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct
     3961 tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcc
     4021 tggaaacggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg aacccgcgct
     4081 atgacggcaa taaaaagaca gaataaaacg cacgggtgtt gggtcgtttg ttcataaacg
     4141 cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc attggggcca
     4201 atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg aaggcccagg
     4261 gctcgcagcc aacgtcgggg cggcaggccc tgccatagca gatctgcgca gctggggctc
     4321 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     4381 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     4441 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg gcatcccttt
     4501 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     4561 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     4621 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     4681 ttcttttgat ttataaggga ttttggggat ttcggcctat tggttaaaaa atgagctgat
     4741 ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag
     4801 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     4861 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     4921 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     4981 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     5041 gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
     5101 tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga
     5161 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     5221 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     5281 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     5341 ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     5401 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     5461 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     5521 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     5581 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     5641 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     5701 cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     5761 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     5821 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     5881 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     5941 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgcgaaa tgaccgacca
     6001 agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt
     6061 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat
     6121 gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag
     6181 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     6241 gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt
     6301 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     6361 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     6421 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     6481 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     6541 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     6601 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     6661 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     6721 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     6781 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     6841 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     6901 gctttctcaa tgctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     6961 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     7021 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     7081 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     7141 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     7201 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     7261 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     7321 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     7381 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     7441 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     7501 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     7561 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     7621 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     7681 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     7741 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     7801 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     7861 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     7921 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     7981 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     8041 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     8101 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     8161 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     8221 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     8281 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     8341 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     8401 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     8461 acctgacgtc gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc
     8521 tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct
     8581 gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg
     8641 aagaatctgc ttagg
//
