LOCUS       V032733                 8927 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V032733
VERSION     V032733
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             2303..2764
                     /label="Id1(NM_001355113)"
                     /note="Id1(NM_001355113)"
                     /gene="Id1"
     CDS             2771..2836
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        2880..3091
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3104..3372
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      3401..3410
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             3431..4093
                     /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             4163..4216
                     /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             4223..5017
                     /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="MAEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*"
                     /label="NeoR/KanR"
     misc_feature    5024..5612
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5684..5917
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    5989..6110
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6129..6264
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6302..6318)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6326..6342
                     /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(6350..6380)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6395..6416
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6704..7292)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7463..8323)
                     /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(8324..8428)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     8902..8918
                     /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 gaattcgcca ccatgaaggt cgccagtggc agtgccgcag ccgctgcagg
     2341 ccctagctgt tcgctgaagg cgggcaggac agcgggcgag gtggtacttg gtctgtcgga
     2401 gcaaagcgtg gccatctcgc gctgcgctgg gacgcgcctg cccgccttgc tggacgagca
     2461 gcaggtgaac gtcctgctct acgacatgaa cggctgctac tcacgcctca aggagctggt
     2521 gcccaccctg ccccagaacc gcaaagtgag caaggtggag atcctgcagc atgtaatcga
     2581 ctacatcagg gacctgcagc tggagctgaa ctcggagtct gaagtcggga ccaccggagg
     2641 ccggggactg cctgtccgcg ccccgctcag caccctgaac ggcgagatca gtgccttggc
     2701 ggccgaggtg aggcggcatg tgttccagcc gacgatcgca tcttgtgtcg ctgaggcggc
     2761 gcacggatcc gactacaaag accatgacgg tgattataaa gatcatgaca tcgactacaa
     2821 ggatgacgat gacaagtagc ggccgcgaag gatctgcgat cgctccggtg cccgtcagtg
     2881 ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg gcaattgaac
     2941 gggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg
     3001 cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct
     3061 ttttcgcaac gggtttgccg ccagaacaca gctgaagctt cgaggggctc gcatctctcc
     3121 ttcacgcgcc cgccgcccta cctgaggccg ccatccacgc cggttgagtc gcgttctgcc
     3181 gcctcccgcc tgtggtgcct cctgaactgc gtccgccgtc taggtaagtt taaagctcag
     3241 gtcgagaccg ggcctttgtc cggcgctccc ttggagccta cctagactca gccggctctc
     3301 cacgctttgc ctgaccctgc ttgctcaact ctacgtcttt gtttcgtttt ctgttctgcg
     3361 ccgttacaga tccaagctgt gaccggcgcc tacgctagac gccaccatgg agagcgacga
     3421 gagcggcctg cccgccatgg agatcgagtg ccgcatcacc ggcaccctga acggcgtgga
     3481 gttcgagctg gtgggcggcg gagagggcac ccccaagcag ggccgcatga ccaacaagat
     3541 gaagagcacc aaaggcgccc tgaccttcag cccctacctg ctgagccacg tgatgggcta
     3601 cggcttctac cacttcggca cctaccccag cggctacgag aaccccttcc tgcacgccat
     3661 caacaacggc ggctacacca acacccgcat cgagaagtac gaggacggcg gcgtgctgca
     3721 cgtgagcttc agctaccgct acgaggccgg ccgcgtgatc ggcgacttca aggtggtggg
     3781 caccggcttc cccgaggaca gcgtgatctt caccgacaag atcatccgca gcaacgccac
     3841 cgtggagcac ctgcacccca tgggcgataa cgtgctggtg ggcagcttcg cccgcacctt
     3901 cagcctgcgc gacggcggct actacagctt cgtggtggac agccacatgc acttcaagag
     3961 cgccatccac cccagcatcc tgcagaacgg gggccccatg ttcgccttcc gccgcgtgga
     4021 ggagctgcac agcaacaccg agctgggcat cgtggagtac cagcacgcct tcaagacccc
     4081 catcgccttc gccagatccc gcgctcagtc gtccaattct gccgtggacg gcaccgccgg
     4141 acccggctcc accggatctc gcgagggcag aggaagtctt ctaacatgcg gtgacgtgga
     4201 ggagaatccc ggccctgcca ccatggctga acaagatgga ttgcacgcag gttctccggc
     4261 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
     4321 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
     4381 gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac
     4441 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     4501 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     4561 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     4621 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     4681 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     4741 gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
     4801 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     4861 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     4921 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     4981 catcgccttc tatcgccttc ttgacgagtt cttctgagtc gacaatcaac ctctggatta
     5041 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg
     5101 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc
     5161 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca
     5221 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac
     5281 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact
     5341 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc
     5401 cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctgtg ttgccacctg
     5461 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc
     5521 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac
     5581 gagtcggatc tccctttggg ccgcctcccc gcggtacctt taagaccaat gacttacaag
     5641 gcagctgtag atcttagcca ctttttaaaa gaaaaggggg gactggaagg gctaattcac
     5701 tcccaacgaa gataagatct gctttttgct tgtactgggt ctctctggtt agaccagatc
     5761 tgagcctggg agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg
     5821 ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc
     5881 ctcagaccct tttagtcagt gtggaaaatc tctagcagta gtagttcatg tcatcttatt
     5941 attcagtatt tataacttgc aaagaaatga atatcagaga gtgagaggaa cttgtttatt
     6001 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     6061 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg
     6121 ctctagctat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     6181 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     6241 agtgaggagg cttttttgga ggcctagact tttgcagaga cggcccaaat tcgtaatcat
     6301 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     6361 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     6421 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     6481 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     6541 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     6601 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     6661 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     6721 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     6781 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     6841 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     6901 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     6961 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     7021 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     7081 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     7141 gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     7201 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     7261 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     7321 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     7381 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     7441 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     7501 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     7561 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     7621 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     7681 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     7741 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     7801 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     7861 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     7921 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     7981 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     8041 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     8101 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     8161 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     8221 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     8281 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     8341 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     8401 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct
     8461 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc
     8521 gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg
     8581 tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg
     8641 gtgttggcgg gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag
     8701 tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc
     8761 gccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc
     8821 tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag
     8881 ggttttccca gtcacgacgt tgtaaaacga cggccagtgc caagctg
//
