LOCUS       V047475                 9391 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V047475
VERSION     V047475
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     protein_bind    113..134
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     promoter        149..179
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    187..203
                     /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"
     primer_bind     211..227
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     promoter        248..266
                     /note="promoter for bacteriophage T3 RNA polymerase"
                     /label="T3 promoter"
     promoter        294..520
                     /note="Rous sarcoma virus enhancer/promoter"
                     /label="RSV promoter"
     LTR             521..701
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
                     /label="5' LTR (truncated)"
     misc_feature    748..873
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
                     /label="HIV-1 Ψ"
     misc_feature    1366..1599
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
                     /label="RRE"
     CDS             1784..1828
                     /codon_start=1
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /product="antigenic peptide corresponding to amino acids
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik
                     et al., 2013)"
                     /transl_table=1
                     /translation="KNEQELLELDKWASL"
                     /label="gp41 peptide"
     misc_feature    2095..2211
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
                     /label="cPPT/CTS"
     enhancer        2234..2537
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        2538..2741
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     regulatory      2818..2827
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             2827..2892
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             2899..3717
                     /label="Bsg(NM_001077184)"
                     /note="Bsg(NM_001077184)"
                     /gene="Bsg"
     CDS             3754..4473
                     /codon_start=1
                     /note="mammalian codon-optimized"
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /transl_table=1
                     /translation="M,V,SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKL
                     TLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFK
                     DDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNG
                     IKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMV
                     LLEFVTAAGITLGMDELYK*"
                     /label="EGFP"
     promoter        4523..4852
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             4862..5461
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /note="confers resistance to puromycin"
                     /product="puromycin N-acetyltransferase"
                     /transl_table=1
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA*"
                     /label="PuroR"
     misc_feature    5528..6116
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6203..6436
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6508..6629
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6669..6804
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     promoter        complement(6825..6843)
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /label="T7 promoter"
     primer_bind     complement(6853..6869)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 fwd"
     rep_origin      7011..7466
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        7492..7596
                     /gene="bla"
                     /label="AmpR promoter"
     CDS             7597..8457
                     /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"
     rep_origin      8628..9216
                     /direction=RIGHT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
ORIGIN
        1 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
       61 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
      121 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
      181 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
      241 agcgcgcaat taaccctcac taaagggaac aaaagctgga gctgcaagct taatgtagtc
      301 ttatgcaata ctcttgtagt cttgcaacat ggtaacgatg agttagcaac atgccttaca
      361 aggagagaaa aagcaccgtg catgccgatt ggtggaagta aggtggtacg atcgtgcctt
      421 attaggaagg caacagacgg gtctgacatg gattggacga accactgaat tgccgcattg
      481 cagagatatt gtatttaagt gcctagctcg atacataaac gggtctctct ggttagacca
      541 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag
      601 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag
      661 atccctcaga cccttttagt cagtgtggaa aatctctagc agtggcgccc gaacagggac
      721 ttgaaagcga aagggaaacc agaggagctc tctcgacgca ggactcggct tgctgaagcg
      781 cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt gactagcgga
      841 ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag aattagatcg
      901 cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt aaaacatata
      961 gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt agaaacatca
     1021 gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg atcagaagaa
     1081 cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag gatagagata
     1141 aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag taagaccacc
     1201 gcacagcaag cggccgctga tcttcagacc tggaggagga gatatgaggg acaattggag
     1261 aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag cacccaccaa
     1321 ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag ctttgttcct
     1381 tgggttcttg ggagcagcag gaagcactat gggcgcagcg tcaatgacgc tgacggtaca
     1441 ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga gggctattga
     1501 ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc aggcaagaat
     1561 cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg gttgctctgg
     1621 aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata aatctctgga
     1681 acagatttgg aatcacacga cctggatgga gtgggacaga gaaattaaca attacacaag
     1741 cttaatacac tccttaattg aagaatcgca aaaccagcaa gaaaagaatg aacaagaatt
     1801 attggaatta gataaatggg caagtttgtg gaattggttt aacataacaa attggctgtg
     1861 gtatataaaa ttattcataa tgatagtagg aggcttggta ggtttaagaa tagtttttgc
     1921 tgtactttct atagtgaata gagttaggca gggatattca ccattatcgt ttcagaccca
     1981 cctcccaacc ccgaggggac ccgacaggcc cgaaggaata gaagaagaag gtggagagag
     2041 agacagagac agatccattc gattagtgaa cggatctcga cggtatcggt taacttttaa
     2101 aagaaaaggg gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac
     2161 agacatacaa actaaagaat tacaaaaaca aattacaaaa ttcaaaattt tatcgataag
     2221 cttgggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
     2281 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
     2341 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
     2401 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
     2461 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
     2521 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
     2581 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
     2641 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
     2701 gcggtaggcg tgtacggtgg gaggtctata taagcagagc tcgtttagtg aaccgtcaga
     2761 tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccga ctctagagcc
     2821 accatggact acaaagacca tgacggtgat tataaagatc atgacatcga ctacaaggat
     2881 gacgatgaca aggatatcat ggcggcggcg ctgctgctgg cgctggcctt cacgctcttg
     2941 agcggccaag gcgcctgcgc ggcggcgggc accatccaaa cctctgtcca ggaagtcaac
     3001 tccaaaacac agcttacctg ctctttgaac agcagtggcg ttgacatcgt tggccaccgc
     3061 tggatgagag gtggcaaggt actgcaggag gacactctgc ccgacctgca tacgaagtac
     3121 atagtggacg cagatgaccg ctctggggaa tattcctgca tcttccttcc tgagcctgtg
     3181 ggcagaagcg agatcaatgt ggaagggcca cccaggatca aggtcggaaa gaaatcagag
     3241 cattccagtg agggagagct tgcgaaactg gtctgcaagt ccgatgcatc ctaccctcct
     3301 attacagatt ggttctggtt taagacctct gacactgggg aagaagaggc aatcaccaat
     3361 agcactgaag ccaatggcaa gtatgtggtg gtatccacgc ctgagaagtc acagctgacc
     3421 atcagcaacc ttgacgtaaa tgttgaccct ggcacctacg tgtgtaatgc caccaacgcc
     3481 cagggcacta ctcgggaaac catctcactg cgtgtgcgga gccgcatggc agccctctgg
     3541 cccttcctag gcatcgtggc tgaggtcctg gtgttggtta ccatcatctt tatctatgag
     3601 aagaggcgga agccagacca gaccctggac gaggatgacc ctggcgccgc cccactgaag
     3661 ggcagtggaa ctcacatgaa tgacaaggac aagaatgtac gccagaggaa cgccaccctc
     3721 gagggcggtg gaggcagtgg tggaggcggt agcatggtga gcaagggcga ggagctgttc
     3781 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
     3841 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
     3901 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg
     3961 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
     4021 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
     4081 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
     4141 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac
     4201 aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc
     4261 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     4321 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc
     4381 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     4441 atcactctcg gcatggacga gctgtacaag taacgggatc cggactagtt aacgcgtcga
     4501 ccagtggtcg accctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     4561 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     4621 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     4681 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     4741 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga
     4801 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagcttac
     4861 catgaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt
     4921 acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga
     4981 ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga
     5041 catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga
     5101 gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg
     5161 ttcccggctg gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga
     5221 gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac caccagggca agggtctggg
     5281 cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct
     5341 ggagacctcc gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc
     5401 cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg
     5461 acgcccgccc cacgacccgc agcgcccgac cgaaaggagc gcacgacccc atgcatcgcc
     5521 agtcgacaat caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta
     5581 tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc
     5641 ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga
     5701 ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac
     5761 ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc
     5821 cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc
     5881 tcggctgttg ggcactgaca attccgtggt gttgtcgggg aagctgacgt cctttccatg
     5941 gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc
     6001 ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc
     6061 gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcctgg
     6121 aattcgagct cggtaccttt aagaccaatg acttacaagg cagctgtaga tcttagccac
     6181 tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag acaagatctg
     6241 ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc
     6301 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
     6361 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
     6421 tggaaaatct ctagcagtag tagttcatgt catcttatta ttcagtattt ataacttgca
     6481 aagaaatgaa tatcagagag tgagaggaac ttgtttattg cagcttataa tggttacaaa
     6541 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     6601 ggtttgtcca aactcatcaa tgtatcttat catgtctggc tctagctatc ccgcccctaa
     6661 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     6721 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     6781 agtgaggagg cttttttgga ggcctaggga cgtacccaat tcgccctata gtgagtcgta
     6841 ttacgcgcgc tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac
     6901 ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc
     6961 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggg acgcgccctg
     7021 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     7081 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     7141 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     7201 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     7261 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     7321 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     7381 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
     7441 taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt gcgcggaacc
     7501 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     7561 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     7621 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     7681 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     7741 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     7801 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     7861 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     7921 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     7981 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     8041 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     8101 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     8161 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     8221 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     8281 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     8341 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     8401 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     8461 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     8521 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     8581 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     8641 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     8701 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     8761 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     8821 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     8881 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     8941 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     9001 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     9061 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     9121 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     9181 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     9241 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     9301 tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg
     9361 accgagcgca gcgagtcagt gagcgaggaa g
//
