LOCUS       V036017                 8857 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V036017
VERSION     V036017
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..2883
                     /label="KRAS(NM_033360)"
                     /note="KRAS(NM_033360)"
                     /gene="KRAS"
     CDS             2890..2955
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        2999..3210
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3223..3491
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      3520..3529
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             3550..4212
                     /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             4282..4335
                     /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             4336..4935
                     /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    4936..5524
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5598..5831
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    5903..6024
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6064..6199
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6232..6248)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6256..6272
                     /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(6280..6310)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6325..6346
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6634..7222)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7393..8253)
                     /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(8254..8358)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     8832..8848
                     /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 gccaccatga ctgaatataa acttgtggta
     2341 gttggagctg gtggcgtagg caagagtgcc ttgacgatac agctaattca gaatcatttt
     2401 gtggacgaat atgatccaac aatagaggat tcctacagga agcaagtagt aattgatgga
     2461 gaaacctgtc tcttggatat tctcgacaca gcaggtcaag aggagtacag tgcaatgagg
     2521 gaccagtaca tgaggactgg ggagggcttt ctttgtgtat ttgccataaa taatactaaa
     2581 tcatttgaag atattcacca ttatagagaa caaattaaaa gagttaagga ctctgaagat
     2641 gtacctatgg tcctagtagg aaataaatgt gatttgcctt ctagaacagt agacacaaaa
     2701 caggctcagg acttagcaag aagttatgga attcctttta ttgaaacatc agcaaagaca
     2761 agacagagag tggaggatgc tttttataca ttggtgagag agatccgaca atacagattg
     2821 aaaaaaatca gcaaagaaga aaagactcct ggctgtgtga aaattaaaaa atgcattata
     2881 atgggatccg actacaaaga ccatgacggt gattataaag atcatgacat cgactacaag
     2941 gatgacgatg acaagtagcg gccgcgaagg atctgcgatc gctccggtgc ccgtcagtgg
     3001 gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg caattgaacg
     3061 ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc
     3121 ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt
     3181 tttcgcaacg ggtttgccgc cagaacacag ctgaagcttc gaggggctcg catctctcct
     3241 tcacgcgccc gccgccctac ctgaggccgc catccacgcc ggttgagtcg cgttctgccg
     3301 cctcccgcct gtggtgcctc ctgaactgcg tccgccgtct aggtaagttt aaagctcagg
     3361 tcgagaccgg gcctttgtcc ggcgctccct tggagcctac ctagactcag ccggctctcc
     3421 acgctttgcc tgaccctgct tgctcaactc tacgtctttg tttcgttttc tgttctgcgc
     3481 cgttacagat ccaagctgtg accggcgcct acgctagacg ccaccatgga gagcgacgag
     3541 agcggcctgc ccgccatgga gatcgagtgc cgcatcaccg gcaccctgaa cggcgtggag
     3601 ttcgagctgg tgggcggcgg agagggcacc cccaagcagg gccgcatgac caacaagatg
     3661 aagagcacca aaggcgccct gaccttcagc ccctacctgc tgagccacgt gatgggctac
     3721 ggcttctacc acttcggcac ctaccccagc ggctacgaga accccttcct gcacgccatc
     3781 aacaacggcg gctacaccaa cacccgcatc gagaagtacg aggacggcgg cgtgctgcac
     3841 gtgagcttca gctaccgcta cgaggccggc cgcgtgatcg gcgacttcaa ggtggtgggc
     3901 accggcttcc ccgaggacag cgtgatcttc accgacaaga tcatccgcag caacgccacc
     3961 gtggagcacc tgcaccccat gggcgataac gtgctggtgg gcagcttcgc ccgcaccttc
     4021 agcctgcgcg acggcggcta ctacagcttc gtggtggaca gccacatgca cttcaagagc
     4081 gccatccacc ccagcatcct gcagaacggg ggccccatgt tcgccttccg ccgcgtggag
     4141 gagctgcaca gcaacaccga gctgggcatc gtggagtacc agcacgcctt caagaccccc
     4201 atcgccttcg ccagatcccg cgctcagtcg tccaattctg ccgtggacgg caccgccgga
     4261 cccggctcca ccggatctcg cgagggcaga ggaagtcttc taacatgcgg tgacgtggag
     4321 gagaatcccg gccctatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
     4381 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
     4441 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
     4501 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc
     4561 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
     4621 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg
     4681 caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag
     4741 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg
     4801 gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc
     4861 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
     4921 aagcccggtg cctgaaatca acctctggat tacaaaattt gtgaaagatt gactggtatt
     4981 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat
     5041 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct
     5101 ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct
     5161 gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc
     5221 gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg
     5281 acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc
     5341 tttccttggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac
     5401 gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg
     5461 cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc
     5521 ccgcctggta cctttaagac caatgactta caaggcagct gtagatctta gccacttttt
     5581 aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagataag atctgctttt
     5641 tgcttgtact gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact
     5701 agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc
     5761 ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa
     5821 aatctctagc agtagtagtt catgtcatct tattattcag tatttataac ttgcaaagaa
     5881 atgaatatca gagagtgaga ggaacttgtt tattgcagct tataatggtt acaaataaag
     5941 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     6001 gtccaaactc atcaatgtat cttatcatgt ctggctctag ctatcccgcc cctaactccg
     6061 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     6121 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     6181 ggaggctttt ttggaggcct agacttttgc agagaccaaa ttcgtaatca tgtcatagct
     6241 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     6301 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     6361 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     6421 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     6481 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     6541 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     6601 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     6661 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     6721 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     6781 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     6841 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     6901 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     6961 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     7021 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     7081 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     7141 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     7201 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     7261 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     7321 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     7381 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     7441 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     7501 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     7561 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     7621 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     7681 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     7741 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     7801 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     7861 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     7921 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     7981 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     8041 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     8101 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     8161 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     8221 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     8281 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     8341 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     8401 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg
     8461 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     8521 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     8581 gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat
     8641 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc
     8701 attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca
     8761 gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca
     8821 gtcacgacgt tgtaaaacga cggccagtgc caagctg
//
