LOCUS       V035525                 8557 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V035525
VERSION     V035525
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..1921
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
                     /label="cPPT/CTS"
     promoter        2011..2214
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     CDS             2317..2583
                     /label="Timm9(NM_013896)"
                     /note="Timm9(NM_013896)"
                     /gene="Timm9"
     CDS             2590..2655
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        2699..2910
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             2923..3191
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      3220..3229
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             3250..3912
                     /codon_start=1
                     /product="green fluorescent protein 2 from Pontellina
                     plumata, also known as ppluGFP2 (Shagin et al., 2004)"
                     /transl_table=1
                     /translation="PAMEIECRITGTLNGVEFELVGGGEGTPKQGRMTNKMKSTKGALT
                     FSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYRY
                     EAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDGG
                     YYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA"
                     /label="CopGFP"
     CDS             3982..4035
                     /codon_start=1
                     /note="Eukaryotic ribosomes fail to insert a peptide bond
                     between the Gly and Pro residues, yielding separate
                     polypeptides."
                     /product="2A peptide from&nbsp;Thosea asigna virus capsid
                     protein"
                     /protein_id=""
                     /transl_table=1
                     /translation="EGRGSLLTCGDVEENPGP"
                     /label="T2A"
     CDS             4036..4635
                     /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    4636..5224
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5298..5531
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    5603..5724
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      5764..5899
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(5932..5948)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    5956..5972
                     /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(5980..6010)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6025..6046
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6334..6922)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7093..7953)
                     /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(7954..8058)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     8532..8548
                     /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 attacaaaaa ttcaaaattt
     1921 tatcgatact agtattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     1981 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     2041 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     2101 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     2161 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggttt atataagcag agctcgttta
     2221 gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagattc
     2281 tagagctagc ctgatacgaa ctcggaattc gccaccatgg ctgcacagat accggaatct
     2341 gaccagataa aacagtttaa ggagttcctg ggaacctaca ataaacttac agaaacctgc
     2401 tttttggact gtgttaaaga cttcacaaca agagaggtga aacctgaaga ggtgacctgt
     2461 tcagaacatt gcttacagaa atatttaaaa atgacacaga ggatatccgt gagatttcag
     2521 gaatatcaca ttcagcagaa tgaagccctg gcagccaaag caggcctcct gggccagcca
     2581 cggggatccg actacaaaga ccatgacggt gattataaag atcatgacat cgactacaag
     2641 gatgacgatg acaagtagcg gccgcgaagg atctgcgatc gctccggtgc ccgtcagtgg
     2701 gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg caattgaacg
     2761 ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc
     2821 ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt
     2881 tttcgcaacg ggtttgccgc cagaacacag ctgaagcttc gaggggctcg catctctcct
     2941 tcacgcgccc gccgccctac ctgaggccgc catccacgcc ggttgagtcg cgttctgccg
     3001 cctcccgcct gtggtgcctc ctgaactgcg tccgccgtct aggtaagttt aaagctcagg
     3061 tcgagaccgg gcctttgtcc ggcgctccct tggagcctac ctagactcag ccggctctcc
     3121 acgctttgcc tgaccctgct tgctcaactc tacgtctttg tttcgttttc tgttctgcgc
     3181 cgttacagat ccaagctgtg accggcgcct acgctagacg ccaccatgga gagcgacgag
     3241 agcggcctgc ccgccatgga gatcgagtgc cgcatcaccg gcaccctgaa cggcgtggag
     3301 ttcgagctgg tgggcggcgg agagggcacc cccaagcagg gccgcatgac caacaagatg
     3361 aagagcacca aaggcgccct gaccttcagc ccctacctgc tgagccacgt gatgggctac
     3421 ggcttctacc acttcggcac ctaccccagc ggctacgaga accccttcct gcacgccatc
     3481 aacaacggcg gctacaccaa cacccgcatc gagaagtacg aggacggcgg cgtgctgcac
     3541 gtgagcttca gctaccgcta cgaggccggc cgcgtgatcg gcgacttcaa ggtggtgggc
     3601 accggcttcc ccgaggacag cgtgatcttc accgacaaga tcatccgcag caacgccacc
     3661 gtggagcacc tgcaccccat gggcgataac gtgctggtgg gcagcttcgc ccgcaccttc
     3721 agcctgcgcg acggcggcta ctacagcttc gtggtggaca gccacatgca cttcaagagc
     3781 gccatccacc ccagcatcct gcagaacggg ggccccatgt tcgccttccg ccgcgtggag
     3841 gagctgcaca gcaacaccga gctgggcatc gtggagtacc agcacgcctt caagaccccc
     3901 atcgccttcg ccagatcccg cgctcagtcg tccaattctg ccgtggacgg caccgccgga
     3961 cccggctcca ccggatctcg cgagggcaga ggaagtcttc taacatgcgg tgacgtggag
     4021 gagaatcccg gccctatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
     4081 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
     4141 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
     4201 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc
     4261 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
     4321 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg
     4381 caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag
     4441 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg
     4501 gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc
     4561 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
     4621 aagcccggtg cctgaaatca acctctggat tacaaaattt gtgaaagatt gactggtatt
     4681 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat
     4741 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct
     4801 ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct
     4861 gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc
     4921 gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg
     4981 acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc
     5041 tttccttggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac
     5101 gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg
     5161 cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc
     5221 ccgcctggta cctttaagac caatgactta caaggcagct gtagatctta gccacttttt
     5281 aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagataag atctgctttt
     5341 tgcttgtact gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact
     5401 agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc
     5461 ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa
     5521 aatctctagc agtagtagtt catgtcatct tattattcag tatttataac ttgcaaagaa
     5581 atgaatatca gagagtgaga ggaacttgtt tattgcagct tataatggtt acaaataaag
     5641 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     5701 gtccaaactc atcaatgtat cttatcatgt ctggctctag ctatcccgcc cctaactccg
     5761 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     5821 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     5881 ggaggctttt ttggaggcct agacttttgc agagaccaaa ttcgtaatca tgtcatagct
     5941 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     6001 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     6061 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     6121 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     6181 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     6241 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     6301 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     6361 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     6421 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     6481 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     6541 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     6601 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     6661 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     6721 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     6781 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     6841 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     6901 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     6961 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     7021 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     7081 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     7141 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     7201 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     7261 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     7321 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     7381 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     7441 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     7501 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     7561 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     7621 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     7681 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     7741 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     7801 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     7861 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     7921 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     7981 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     8041 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     8101 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg
     8161 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     8221 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     8281 gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat
     8341 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc
     8401 attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca
     8461 gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca
     8521 gtcacgacgt tgtaaaacga cggccagtgc caagctg
//
