LOCUS       V035921                 8598 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V035921
VERSION     V035921
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..1466
                     /codon_start=1
                     /note="mammalian codon-optimized"
                     /product="monomeric derivative of red fluorescent protein
                     from Entacmaea quadricolor&nbsp;(Merzlyak et al., 2007)"
                     /transl_table=1
                     /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFMYGSRTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEANTEMLYPADGGLEGRSD
                     MALKLVGGGHLICNFKTTYRSKKPAKNLKMPGVYYVDHRLERIKEADKETYVEQHEVAV
                     ARYCDLPSKLGHKLN"
                     /label="TagRFP"
     CDS             1503..3278
                     /label="TGFBR2(NM_001024847)"
                     /note="TGFBR2(NM_001024847)"
                     /gene="TGFBR2"
     CDS             3285..3350
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    3374..3962
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    4028..4076
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      4278..4706
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        4720..5049
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             5116..5910
                     /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    6086..6207
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(6256..6272)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6280..6296
                     /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(6304..6334)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6349..6370
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6658..7246)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7417..8277)
                     /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(8278..8382)
                     /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 ccaccatggt gtctaagggc gaagagctga
      781 ttaaggagaa catgcacatg aagctgtaca tggagggcac cgtgaacaac caccacttca
      841 agtgcacatc cgagggcgaa ggcaagccct acgagggcac ccagaccatg agaatcaagg
      901 tggtcgaggg cggccctctc cccttcgcct tcgacatcct ggctaccagc ttcatgtacg
      961 gcagcagaac cttcatcaac cacacccagg gcatccccga cttctttaag cagtccttcc
     1021 ctgagggctt cacatgggag agagtcacca catacgaaga cgggggcgtg ctgaccgcta
     1081 cccaggacac cagcctccag gacggctgcc tcatctacaa cgtcaagatc agaggggtga
     1141 acttcccatc caacggccct gtgatgcaga agaaaacact cggctgggag gccaacaccg
     1201 agatgctgta ccccgctgac ggcggcctgg aaggcagaag cgacatggcc ctgaagctcg
     1261 tgggcggggg ccacctgatc tgcaacttca agaccacata cagatccaag aaacccgcta
     1321 agaacctcaa gatgcccggc gtctactatg tggaccacag actggaaaga atcaaggagg
     1381 ccgacaaaga gacctacgtc gagcagcacg aggtggctgt ggccagatac tgcgacctcc
     1441 ctagcaaact ggggcacaaa cttaatgaat tcggtggagg cggatcagga ggcggtggat
     1501 ctatgggtcg ggggctgctc aggggcctgt ggccgctgca catcgtcctg tggacgcgta
     1561 tcgccagcac gatcccaccg cacgttcaga agtcggatgt ggaaatggag gcccagaaag
     1621 atgaaatcat ctgccccagc tgtaatagga ctgcccatcc actgagacat attaataacg
     1681 acatgatagt cactgacaac aacggtgcag tcaagtttcc acaactgtgt aaattttgtg
     1741 atgtgagatt ttccacctgt gacaaccaga aatcctgcat gagcaactgc agcatcacct
     1801 ccatctgtga gaagccacag gaagtctgtg tggctgtatg gagaaagaat gacgagaaca
     1861 taacactaga gacagtttgc catgacccca agctccccta ccatgacttt attctggaag
     1921 atgctgcttc tccaaagtgc attatgaagg aaaaaaaaaa gcctggtgag actttcttca
     1981 tgtgttcctg tagctctgat gagtgcaatg acaacatcat cttctcagaa gaatataaca
     2041 ccagcaatcc tgacttgttg ctagtcatat ttcaagtgac aggcatcagc ctcctgccac
     2101 cactgggagt tgccatatct gtcatcatca tcttctactg ctaccgcgtt aaccggcagc
     2161 agaagctgag ttcaacctgg gaaaccggca agacgcggaa gctcatggag ttcagcgagc
     2221 actgtgccat catcctggaa gatgaccgct ctgacatcag ctccacgtgt gccaacaaca
     2281 tcaaccacaa cacagagctg ctgcccattg agctggacac cctggtgggg aaaggtcgct
     2341 ttgctgaggt ctataaggcc aagctgaagc agaacacttc agagcagttt gagacagtgg
     2401 cagtcaagat ctttccctat gaggagtatg cctcttggaa gacagagaag gacatcttct
     2461 cagacatcaa tctgaagcat gagaacatac tccagttcct gacggctgag gagcggaaga
     2521 cggagttggg gaaacaatac tggctgatca ccgccttcca cgccaagggc aacctacagg
     2581 agtacctgac gcggcatgtc atcagctggg aggacctgcg caagctgggc agctccctcg
     2641 cccgggggat tgctcacctc cacagtgatc acactccatg tgggaggccc aagatgccca
     2701 tcgtgcacag ggacctcaag agctccaata tcctcgtgaa gaacgaccta acctgctgcc
     2761 tgtgtgactt tgggctttcc ctgcgtctgg accctactct gtctgtggat gacctggcta
     2821 acagtgggca ggtgggaact gcaagataca tggctccaga agtcctagaa tccaggatga
     2881 atttggagaa tgttgagtcc ttcaagcaga ccgatgtcta ctccatggct ctggtgctct
     2941 gggaaatgac atctcgctgt aatgcagtgg gagaagtaaa agattatgag cctccatttg
     3001 gttccaaggt gcgggagcac ccctgtgtcg aaagcatgaa ggacaacgtg ttgagagatc
     3061 gagggcgacc agaaattccc agcttctggc tcaaccacca gggcatccag atggtgtgtg
     3121 agacgttgac tgagtgctgg gaccacgacc cagaggcccg tctcacagcc cagtgtgtgg
     3181 cagaacgctt cagtgagctg gagcatctgg acaggctctc ggggaggagc tgctcggagg
     3241 agaagattcc tgaagacggc tccctaaaca ctaccaaagg atccgactac aaagaccatg
     3301 acggtgatta taaagatcat gacatcgact acaaggatga cgatgacaag tagtgagggc
     3361 ccgatatctc gacaatcaac ctctggatta caaaatttgt gaaagattga ctggtattct
     3421 taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc
     3481 tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct
     3541 ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga
     3601 cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc
     3661 tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac
     3721 aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaagc tgacgtcctt
     3781 tccatggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt
     3841 cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc
     3901 tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc
     3961 gcctggaaac gggggaggct aactgaaaca cggaaggaga caataccgga aggaacccgc
     4021 gctatgacgg caataaaaag acagaataaa acgcacgggt gttgggtcgt ttgttcataa
     4081 acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg
     4141 ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc
     4201 agggctcgca gccaacgtcg gggcggcagg ccctgccata gcagatctgc gcagctgggg
     4261 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     4321 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     4381 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc ggggcatccc
     4441 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     4501 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     4561 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     4621 ctattctttt gatttataag ggattttggg gatttcggcc tattggttaa aaaatgagct
     4681 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
     4741 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     4801 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     4861 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     4921 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     4981 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     5041 ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca agagacagga
     5101 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     5161 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     5221 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     5281 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     5341 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     5401 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     5461 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     5521 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     5581 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     5641 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     5701 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     5761 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     5821 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     5881 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgcg aaatgaccga
     5941 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     6001 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     6061 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     6121 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     6181 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     6241 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     6301 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     6361 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     6421 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     6481 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     6541 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     6601 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     6661 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     6721 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     6781 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     6841 ggcgctttct caatgctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     6901 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     6961 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     7021 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     7081 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     7141 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     7201 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     7261 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     7321 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     7381 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     7441 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     7501 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     7561 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     7621 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     7681 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     7741 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     7801 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     7861 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     7921 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     7981 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     8041 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     8101 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     8161 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     8221 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     8281 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     8341 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     8401 gccacctgac gtcgacggat cgggagatct cccgatcccc tatggtcgac tctcagtaca
     8461 atctgctctg atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc
     8521 gctgagtagt gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc
     8581 atgaagaatc tgcttagg
//
