LOCUS       V045448                 8846 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V045448
VERSION     V045448
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     LTR             234..414
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
                     /label="5' LTR (truncated)"
     misc_feature    458..583
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
                     /label="HIV-1 Ψ"
     misc_feature    1076..1309
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
                     /label="RRE"
     CDS             1493..1537
                     /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    1804..1920
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
                     /label="cPPT/CTS"
     promoter        2010..2213
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     CDS             2302..3018
                     /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"
     CDS             3055..3699
                     /label="HMGB1(NM_001313893)"
                     /note="HMGB1(NM_001313893)"
                     /gene="HMGB1"
     CDS             3706..3771
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3816..4027
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             4040..4308
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     CDS             4335..4934
                     /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    4941..5529
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5603..5836
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    5908..6029
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6048..6183
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6221..6237)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6245..6261
                     /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(6269..6299)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6314..6335
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6623..7211)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7382..8242)
                     /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(8243..8347)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     8821..8837
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 fwd"
ORIGIN
        1 acgcgtgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga tgagttagca
       61 acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag taaggtggta
      121 cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac gaaccactga
      181 attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacaata aacgggtctc
      241 tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta
      301 agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact
      361 ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtggcg
      421 cccgaacagg gacctgaaag cgaaagggaa accagagctc tctcgacgca ggactcggct
      481 tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt
      541 gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag
      601 aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt
      661 aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt
      721 agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg
      781 atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag
      841 gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag
      901 taagaccacc gcacagcaag cggccactga tcttcagacc tggaggagga gatatgaggg
      961 acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag
     1021 cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag
     1081 ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcc tcaatgacgc
     1141 tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga
     1201 gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc
     1261 aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg
     1321 gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata
     1381 aatctctgga acagattgga atcacacgac ctggatggag tgggacagag aaattaacaa
     1441 ttacacaagc ttaatacact ccttaattga agaatcgcaa aaccagcaag aaaagaatga
     1501 acaagaatta ttggaattag ataaatgggc aagtttgtgg aattggttta acataacaaa
     1561 ttggctgtgg tatataaaat tattcataat gatagtagga ggcttggtag gtttaagaat
     1621 agtttttgct gtactttcta tagtgaatag agttaggcag ggatattcac cattatcgtt
     1681 tcagacccac ctcccaaccc cgaggggacc cgacaggccc gaaggaatag aagaagaagg
     1741 tggagagaga gacagagaca gatccattcg attagtgaac ggatctcgac ggtatcggtt
     1801 aacttttaaa agaaaagggg ggattggggg gtacagtgca ggggaaagaa tagtagacat
     1861 aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaat tcaaaatttt
     1921 atcgatacta gtattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
     1981 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
     2041 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
     2101 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
     2161 tgacgcaaat gggcggtagg cgtgtacggt gggaggttta tataagcaga gctcgtttag
     2221 tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagattct
     2281 agagctagcg aattcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg
     2341 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag
     2401 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
     2461 ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc
     2521 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     2581 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     2641 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     2701 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc
     2761 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag
     2821 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     2881 gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac
     2941 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     3001 atggacgagc tgtacaaggg cggcggaggg agtggaggag ggggaagtgg atccatgggc
     3061 aaaggagatc ctaagaagcc gagaggcaaa atgtcatcat atgcattttt tgtgcaaact
     3121 tgtcgggagg agcataagaa gaagcaccca gatgcttcag tcaacttctc agagttttct
     3181 aagaagtgct cagagaggtg gaagaccatg tctgctaaag agaaaggaaa atttgaagat
     3241 atggcaaaag cggacaaggc ccgttatgaa agagaaatga aaacctatat ccctcccaaa
     3301 ggggagacaa aaaagaagtt caaggatccc aatgcaccca agaggcctcc ttcggccttc
     3361 ttcctcttct gctctgagta tcgcccaaaa atcaaaggag aacatcctgg cctgtccatt
     3421 ggtgatgttg cgaagaaact gggagagatg tggaataaca ctgctgcaga tgacaagcag
     3481 ccttatgaaa agaaggctgc gaagctgaag gaaaaatacg aaaaggatat tgctgcatat
     3541 cgagctaaag gaaagcctga tgcagcaaaa aagggagttg tcaaggctga aaaaagcaag
     3601 aaaaagaagg aagaggagga agatgaggaa gatgaagagg atgaggagga ggaggaagat
     3661 gaagaagatg aagatgaaga agaagatgat gatgatgaag gatccgacta caaagaccat
     3721 gacggtgatt ataaagatca tgacatcgac tacaaggatg acgatgacaa gtaggcggcc
     3781 gcgaaggatc tgcgatcgct ccggtgcccg tcagtgggca gagcgcacat cgcccacagt
     3841 ccccgagaag ttggggggag gggtcggcaa ttgaacgggt gcctagagaa ggtggcgcgg
     3901 ggtaaactgg gaaagtgatg tcgtgtactg gctccgcctt tttcccgagg gtgggggaga
     3961 accgtatata agtgcagtag tcgccgtgaa cgttcttttt cgcaacgggt ttgccgccag
     4021 aacacagctg aagcttcgag gggctcgcat ctctccttca cgcgcccgcc gccctacctg
     4081 aggccgccat ccacgccggt tgagtcgcgt tctgccgcct cccgcctgtg gtgcctcctg
     4141 aactgcgtcc gccgtctagg taagtttaaa gctcaggtcg agaccgggcc tttgtccggc
     4201 gctcccttgg agcctaccta gactcagccg gctctccacg ctttgcctga ccctgcttgc
     4261 tcaactctac gtctttgttt cgttttctgt tctgcgccgt tacagatcca agctgtgacc
     4321 ggcgcctacg ctagatgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg
     4381 tcccccgggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca
     4441 ccgtcgaccc ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc
     4501 gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct
     4561 ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg
     4621 ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc
     4681 accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg
     4741 gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg
     4801 tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag cggctcggct
     4861 tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca
     4921 agcccggtgc ctgagtcgac aatcaacctc tggattacaa aatttgtgaa agattgactg
     4981 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt
     5041 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc
     5101 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt
     5161 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga
     5221 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct
     5281 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat
     5341 cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct
     5401 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc
     5461 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg
     5521 cctccccgcc tggtaccttt aagaccaatg acttacaagg cagctgtaga tcttagccac
     5581 tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaaa ataagatctg
     5641 ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc
     5701 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
     5761 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
     5821 tggaaaatct ctagcagtag tagttcatgt catcttatta ttcagtattt ataacttgca
     5881 aagaaatgaa tatcagagag tgagaggaac ttgtttattg cagcttataa tggttacaaa
     5941 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     6001 ggtttgtcca aactcatcaa tgtatcttat catgtctggc tctagctatc ccgcccctaa
     6061 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     6121 aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     6181 gcctagactt ttgcagagac ggcccaaatt cgtaatcatg gtcatagctg tttcctgtgt
     6241 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     6301 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     6361 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     6421 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     6481 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     6541 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     6601 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     6661 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     6721 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     6781 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     6841 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     6901 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     6961 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     7021 cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct
     7081 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     7141 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     7201 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     7261 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     7321 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     7381 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     7441 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     7501 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     7561 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     7621 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     7681 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     7741 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     7801 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     7861 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     7921 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     7981 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     8041 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     8101 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     8161 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     8221 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     8281 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     8341 ttccgcgcac atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga
     8401 cattaaccta taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg
     8461 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg
     8521 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct
     8581 ggcttaacta tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa
     8641 taccgcacag atgcgtaagg agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc
     8701 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
     8761 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
     8821 gtaaaacgac ggccagtgcc aagctg
//
