LOCUS       V035526                 9466 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V035526
VERSION     V035526
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..3492
                     /label="Eeig1(NM_153560)"
                     /note="Eeig1(NM_153560)"
                     /gene="Eeig1"
     CDS             3499..3564
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3608..3819
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3832..4100
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      4129..4138
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             4159..4821
                     /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             4891..4944
                     /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             4945..5544
                     /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    5545..6133
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6207..6440
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6512..6633
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6673..6808
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6841..6857)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6865..6881
                     /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(6889..6919)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6934..6955
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7243..7831)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8002..8862)
                     /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(8863..8967)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9441..9457
                     /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 cttttttgat gaagaagaag
     2341 aaatttaaat ttcaaaccac tttcaccttg gaggagctga ctgcggtccc cttcgtgaac
     2401 ggggtcctct tttgcaaggt ccggctgctg gatggcgggg attttgtcag cttgtcgtcc
     2461 agggaggagg tgcaggagaa ctgtgtacga tggcggaaga ggttcacctt tgtgtgtaaa
     2521 atgagtgcca acccggccac aggcctgctg gacccctgca tctttcgagt gtctgtgcgc
     2581 aaggagctga aaggtgggaa ggcgtattcc aagctgggct tcactgactt gaacctagcc
     2641 gagtttgcag gctcaggctc caccgtccgc tgctgcctat tggaaggata cgacaccaag
     2701 aatacccgcc aggacaactc catcctcaag gtcaccattg gcatgttcct gctctctggg
     2761 gacccctgtt tcaagacgcc accgtctacc gccaaatcca tctccatccc gggccaggac
     2821 tcctccctgc agctgacgtg taaaggtggt gggaccagca gcggtggcag cagcagcacc
     2881 aattccctaa ctggatcccg gccccccaag acccggccca ccatcctggg ctcaggacta
     2941 ccagaggagc cagaccagag cctgtccagt cctgaggagg tgttccactc tggccactcc
     3001 cgcaactcca gctatgccag ccagcagtcc aagctatctg gctacagtac agagcactct
     3061 cgctcctcca gcctgtctga cttgacacac cgcagaaata catccaccag cagcagcgcc
     3121 tctggtggcc tcagtatggc tgtagagggt cctgagggca tggagaggga gcatcggccc
     3181 tctgagaagc cacctcggcc tcctgagaag cccccacgcc ccccacgccc cttgcatcta
     3241 tcagatcgct cttttcggcg aaagaaagac tctgtggaga gccacccaac ctgggtagat
     3301 gacactcgaa ttgatgcaga cgacattgtg gagaagataa tgcagagcca ggacttcact
     3361 gatggcagca acactgagga cagtaacctt cggctgttcg tgagccgtga tggctccacc
     3421 acactgagcg gcattcagct gggcaacaga gtctcctcgg gagtctacga gccagtcgta
     3481 attgagagcc acggatccga ctacaaagac catgacggtg attataaaga tcatgacatc
     3541 gactacaagg atgacgatga caagtagcgg ccgcgaagga tctgcgatcg ctccggtgcc
     3601 cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc
     3661 aattgaacgg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac
     3721 tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg
     3781 aacgttcttt ttcgcaacgg gtttgccgcc agaacacagc tgaagcttcg aggggctcgc
     3841 atctctcctt cacgcgcccg ccgccctacc tgaggccgcc atccacgccg gttgagtcgc
     3901 gttctgccgc ctcccgcctg tggtgcctcc tgaactgcgt ccgccgtcta ggtaagttta
     3961 aagctcaggt cgagaccggg cctttgtccg gcgctccctt ggagcctacc tagactcagc
     4021 cggctctcca cgctttgcct gaccctgctt gctcaactct acgtctttgt ttcgttttct
     4081 gttctgcgcc gttacagatc caagctgtga ccggcgccta cgctagacgc caccatggag
     4141 agcgacgaga gcggcctgcc cgccatggag atcgagtgcc gcatcaccgg caccctgaac
     4201 ggcgtggagt tcgagctggt gggcggcgga gagggcaccc ccaagcaggg ccgcatgacc
     4261 aacaagatga agagcaccaa aggcgccctg accttcagcc cctacctgct gagccacgtg
     4321 atgggctacg gcttctacca cttcggcacc taccccagcg gctacgagaa ccccttcctg
     4381 cacgccatca acaacggcgg ctacaccaac acccgcatcg agaagtacga ggacggcggc
     4441 gtgctgcacg tgagcttcag ctaccgctac gaggccggcc gcgtgatcgg cgacttcaag
     4501 gtggtgggca ccggcttccc cgaggacagc gtgatcttca ccgacaagat catccgcagc
     4561 aacgccaccg tggagcacct gcaccccatg ggcgataacg tgctggtggg cagcttcgcc
     4621 cgcaccttca gcctgcgcga cggcggctac tacagcttcg tggtggacag ccacatgcac
     4681 ttcaagagcg ccatccaccc cagcatcctg cagaacgggg gccccatgtt cgccttccgc
     4741 cgcgtggagg agctgcacag caacaccgag ctgggcatcg tggagtacca gcacgccttc
     4801 aagaccccca tcgccttcgc cagatcccgc gctcagtcgt ccaattctgc cgtggacggc
     4861 accgccggac ccggctccac cggatctcgc gagggcagag gaagtcttct aacatgcggt
     4921 gacgtggagg agaatcccgg ccctatgacc gagtacaagc ccacggtgcg cctcgccacc
     4981 cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc
     5041 acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct gcaagaactc
     5101 ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg
     5161 gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc
     5221 ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc
     5281 ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc
     5341 gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag
     5401 cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag
     5461 cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc
     5521 atgacccgca agcccggtgc ctgaaatcaa cctctggatt acaaaatttg tgaaagattg
     5581 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     5641 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     5701 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     5761 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     5821 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     5881 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa
     5941 tcatcgtcct ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc
     6001 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     6061 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg
     6121 gccgcctccc cgcctggtac ctttaagacc aatgacttac aaggcagctg tagatcttag
     6181 ccacttttta aaagaaaagg ggggactgga agggctaatt cactcccaac gaagataaga
     6241 tctgcttttt gcttgtactg ggtctctctg gttagaccag atctgagcct gggagctctc
     6301 tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt
     6361 agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc
     6421 agtgtggaaa atctctagca gtagtagttc atgtcatctt attattcagt atttataact
     6481 tgcaaagaaa tgaatatcag agagtgagag gaacttgttt attgcagctt ataatggtta
     6541 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     6601 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggctctagc tatcccgccc
     6661 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     6721 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     6781 aagtagtgag gaggcttttt tggaggccta gacttttgca gagaccaaat tcgtaatcat
     6841 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     6901 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     6961 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     7021 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     7081 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     7141 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     7201 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     7261 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     7321 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     7381 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     7441 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     7501 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     7561 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     7621 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     7681 aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     7741 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     7801 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     7861 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     7921 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     7981 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     8041 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     8101 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     8161 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     8221 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     8281 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     8341 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     8401 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     8461 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     8521 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     8581 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     8641 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     8701 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     8761 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     8821 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     8881 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     8941 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta
     9001 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg
     9061 tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt
     9121 cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg
     9181 tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg tactgagagt
     9241 gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg
     9301 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     9361 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
     9421 gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc aagctg
//
