LOCUS       V033731                 9136 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V033731
VERSION     V033731
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..3162
                     /label="Hes1(NM_008235)"
                     /note="Hes1(NM_008235)"
                     /gene="Hes1"
     CDS             3169..3234
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3278..3489
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3502..3770
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      3799..3808
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             3829..4491
                     /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             4561..4614
                     /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             4615..5214
                     /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    5215..5803
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5877..6110
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6182..6303
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6343..6478
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6511..6527)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6535..6551
                     /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(6559..6589)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6604..6625
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6913..7501)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7672..8532)
                     /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(8533..8637)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9111..9127
                     /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 gccaccatgc cagctgatat aatggagaaa
     2341 aattcctcct ccccggtggc tgctacccca gccagtgtca acacgacacc ggacaaacca
     2401 aagacggcct ctgagcacag aaagtcatca aagcctatca tggagaagag gcgaagggca
     2461 agaataaatg aaagtctaag ccaactgaaa acactgattt tggatgcact taagaaagat
     2521 agctcccggc attccaagct agagaaggca gacattctgg aaatgactgt gaagcacctc
     2581 cggaacctgc agcgggcgca gatgaccgcc gcgctcagca cagacccgag cgtgttgggg
     2641 aaataccgcg ccggcttcag cgagtgcatg aacgaggtga cccgcttcct gtccacgtgt
     2701 gagggcgtta acaccgaggt gcgcactcgg ctgctgggcc acctggccaa ctgcatgacc
     2761 cagatcaacg ccatgaccta ccccgggcag gcgcaccccg ccttgcaggc gccgccgccg
     2821 ccgcccccgt caggacctgc cggtccccag cacgcgccat tcgcgccgcc gccgccgccg
     2881 cttgtgccca tccccggggg cgcggcgccc cctcccggca gcgcaccctg caagttgggc
     2941 agccaggctg gagaggctgc caaggttttt ggcggcttcc aagtggtgcc ggctcctgac
     3001 ggccaatttg cctttctcat ccccaacggg gccttcgctc acagcggccc ggtcatcccg
     3061 gtctacacca gcaacagtgg gacctcggtg ggtcctaacg cagtgtcacc ttccagtggc
     3121 tcctcgctca cttcggactc catgtggaga ccgtggcgga acggatccga ctacaaagac
     3181 catgacggtg attataaaga tcatgacatc gactacaagg atgacgatga caagtagcgg
     3241 ccgcgaagga tctgcgatcg ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca
     3301 gtccccgaga agttgggggg aggggtcggc aattgaacgg gtgcctagag aaggtggcgc
     3361 ggggtaaact gggaaagtga tgtcgtgtac tggctccgcc tttttcccga gggtggggga
     3421 gaaccgtata taagtgcagt agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc
     3481 agaacacagc tgaagcttcg aggggctcgc atctctcctt cacgcgcccg ccgccctacc
     3541 tgaggccgcc atccacgccg gttgagtcgc gttctgccgc ctcccgcctg tggtgcctcc
     3601 tgaactgcgt ccgccgtcta ggtaagttta aagctcaggt cgagaccggg cctttgtccg
     3661 gcgctccctt ggagcctacc tagactcagc cggctctcca cgctttgcct gaccctgctt
     3721 gctcaactct acgtctttgt ttcgttttct gttctgcgcc gttacagatc caagctgtga
     3781 ccggcgccta cgctagacgc caccatggag agcgacgaga gcggcctgcc cgccatggag
     3841 atcgagtgcc gcatcaccgg caccctgaac ggcgtggagt tcgagctggt gggcggcgga
     3901 gagggcaccc ccaagcaggg ccgcatgacc aacaagatga agagcaccaa aggcgccctg
     3961 accttcagcc cctacctgct gagccacgtg atgggctacg gcttctacca cttcggcacc
     4021 taccccagcg gctacgagaa ccccttcctg cacgccatca acaacggcgg ctacaccaac
     4081 acccgcatcg agaagtacga ggacggcggc gtgctgcacg tgagcttcag ctaccgctac
     4141 gaggccggcc gcgtgatcgg cgacttcaag gtggtgggca ccggcttccc cgaggacagc
     4201 gtgatcttca ccgacaagat catccgcagc aacgccaccg tggagcacct gcaccccatg
     4261 ggcgataacg tgctggtggg cagcttcgcc cgcaccttca gcctgcgcga cggcggctac
     4321 tacagcttcg tggtggacag ccacatgcac ttcaagagcg ccatccaccc cagcatcctg
     4381 cagaacgggg gccccatgtt cgccttccgc cgcgtggagg agctgcacag caacaccgag
     4441 ctgggcatcg tggagtacca gcacgccttc aagaccccca tcgccttcgc cagatcccgc
     4501 gctcagtcgt ccaattctgc cgtggacggc accgccggac ccggctccac cggatctcgc
     4561 gagggcagag gaagtcttct aacatgcggt gacgtggagg agaatcccgg ccctatgacc
     4621 gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tccccagggc cgtacgcacc
     4681 ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc ggaccgccac
     4741 atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc
     4801 aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc
     4861 gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg
     4921 ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg
     4981 tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc
     5041 gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt cctggagacc
     5101 tccgcgcccc gcaacctccc cttctacgag cggctcggct tcaccgtcac cgccgacgtc
     5161 gaggtgcccg aaggaccgcg cacctggtgc atgacccgca agcccggtgc ctgaaatcaa
     5221 cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt
     5281 acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct
     5341 ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc
     5401 gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg
     5461 ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc
     5521 acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc
     5581 actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctgt
     5641 gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca
     5701 gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt
     5761 cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcctggtac ctttaagacc
     5821 aatgacttac aaggcagctg tagatcttag ccacttttta aaagaaaagg ggggactgga
     5881 agggctaatt cactcccaac gaagataaga tctgcttttt gcttgtactg ggtctctctg
     5941 gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc
     6001 tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg
     6061 taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca gtagtagttc
     6121 atgtcatctt attattcagt atttataact tgcaaagaaa tgaatatcag agagtgagag
     6181 gaacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     6241 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     6301 ttatcatgtc tggctctagc tatcccgccc ctaactccgc ccatcccgcc cctaactccg
     6361 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     6421 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     6481 gacttttgca gagaccaaat tcgtaatcat gtcatagctg tttcctgtgt gaaattgtta
     6541 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     6601 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     6661 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     6721 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     6781 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     6841 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     6901 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     6961 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     7021 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     7081 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     7141 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     7201 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     7261 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     7321 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     7381 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     7441 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     7501 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     7561 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     7621 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     7681 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     7741 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     7801 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     7861 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     7921 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     7981 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     8041 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     8101 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     8161 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     8221 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     8281 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     8341 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     8401 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     8461 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     8521 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     8581 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     8641 atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta
     8701 taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa
     8761 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     8821 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta
     8881 tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag
     8941 atgcgtaagg agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc gcaactgttg
     9001 ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc
     9061 tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac
     9121 ggccagtgcc aagctg
//
