LOCUS       V033469                 9481 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V033469
VERSION     V033469
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..3507
                     /label="Ctse(NM_007799)"
                     /note="Ctse(NM_007799)"
                     /gene="Ctse"
     CDS             3514..3579
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3623..3834
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3847..4115
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      4144..4153
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             4174..4836
                     /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             4906..4959
                     /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             4960..5559
                     /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    5560..6148
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6222..6455
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6527..6648
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6688..6823
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6856..6872)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6880..6896
                     /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(6904..6934)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6949..6970
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7258..7846)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8017..8877)
                     /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(8878..8982)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9456..9472
                     /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 agcccctcct tgtgctgctg
     2341 ctgctgctgc tcctggatct ggctcaggcc caaggtgctc tgcacagagt gcccctcaga
     2401 agacatcagt cccttcggaa gaaactacgg gcccaaggac agctctcaga attctggagg
     2461 tctcataact tggacatgac ccgactcagc gagtcctgta atgtgtattc gagtgtcaat
     2521 gaacccctca tcaactacct ggatatggaa tactttggca ccatctccat cggcaccccg
     2581 ccgcagaact tcactgtcat ctttgacacc ggttcatcca acctctgggt cccttctgtg
     2641 tactgcacca gcccagcatg caaggcacac ccagtattcc atccatcgca gtccgacaca
     2701 tacacggagg tagggaatca tttctccatc cagtatggta ccgggagcct gacaggaatc
     2761 attggagctg atcaagtctc tgtggaaggg ttgactgtgg atggccagca gtttggagaa
     2821 agtgtcaagg agccaggcca gacctttgtg aatgcagagt ttgatgggat tctgggtctg
     2881 ggatacccct cattggctgc tggaggagtg accccagtgt ttgacaacat gatggcccag
     2941 aaccttgtgg ctctgcctat gttttctgtc tacttgagca gtgaccctca aggtggctca
     3001 ggcagtgagc tgactttcgg aggctatgac ccctctcatt tctctgggag cctcaactgg
     3061 atcccagtca ccaagcaagc ctattggcag attgccctgg atggaatcca ggtgggagac
     3121 actgtgatgt tctgctccga aggctgccag gccatagtgg acacagggac ctctctcatc
     3181 actggccccc ccgacaagat caaacagctt caagaggcca ttggggccac acccattgat
     3241 ggagaatatg cagtggattg tgccactctc gacacgatgc caaacgttac cttcctcatc
     3301 aacgaggttt catataccct caacccaact gactacatcc tgccggactt ggtggaagga
     3361 atgcagttct gtggcagtgg ctttcaagga cttgacattc cacctccagc tgggcccctc
     3421 tggatcctgg gggatgtctt catccgacag ttctactcag tctttgaccg tggaaataac
     3481 caagtgggat tggcccccgc agttcccgga tccgactaca aagaccatga cggtgattat
     3541 aaagatcatg acatcgacta caaggatgac gatgacaagt agcggccgcg aaggatctgc
     3601 gatcgctccg gtgcccgtca gtgggcagag cgcacatcgc ccacagtccc cgagaagttg
     3661 gggggagggg tcggcaattg aacgggtgcc tagagaaggt ggcgcggggt aaactgggaa
     3721 agtgatgtcg tgtactggct ccgccttttt cccgagggtg ggggagaacc gtatataagt
     3781 gcagtagtcg ccgtgaacgt tctttttcgc aacgggtttg ccgccagaac acagctgaag
     3841 cttcgagggg ctcgcatctc tccttcacgc gcccgccgcc ctacctgagg ccgccatcca
     3901 cgccggttga gtcgcgttct gccgcctccc gcctgtggtg cctcctgaac tgcgtccgcc
     3961 gtctaggtaa gtttaaagct caggtcgaga ccgggccttt gtccggcgct cccttggagc
     4021 ctacctagac tcagccggct ctccacgctt tgcctgaccc tgcttgctca actctacgtc
     4081 tttgtttcgt tttctgttct gcgccgttac agatccaagc tgtgaccggc gcctacgcta
     4141 gacgccacca tggagagcga cgagagcggc ctgcccgcca tggagatcga gtgccgcatc
     4201 accggcaccc tgaacggcgt ggagttcgag ctggtgggcg gcggagaggg cacccccaag
     4261 cagggccgca tgaccaacaa gatgaagagc accaaaggcg ccctgacctt cagcccctac
     4321 ctgctgagcc acgtgatggg ctacggcttc taccacttcg gcacctaccc cagcggctac
     4381 gagaacccct tcctgcacgc catcaacaac ggcggctaca ccaacacccg catcgagaag
     4441 tacgaggacg gcggcgtgct gcacgtgagc ttcagctacc gctacgaggc cggccgcgtg
     4501 atcggcgact tcaaggtggt gggcaccggc ttccccgagg acagcgtgat cttcaccgac
     4561 aagatcatcc gcagcaacgc caccgtggag cacctgcacc ccatgggcga taacgtgctg
     4621 gtgggcagct tcgcccgcac cttcagcctg cgcgacggcg gctactacag cttcgtggtg
     4681 gacagccaca tgcacttcaa gagcgccatc caccccagca tcctgcagaa cgggggcccc
     4741 atgttcgcct tccgccgcgt ggaggagctg cacagcaaca ccgagctggg catcgtggag
     4801 taccagcacg ccttcaagac ccccatcgcc ttcgccagat cccgcgctca gtcgtccaat
     4861 tctgccgtgg acggcaccgc cggacccggc tccaccggat ctcgcgaggg cagaggaagt
     4921 cttctaacat gcggtgacgt ggaggagaat cccggcccta tgaccgagta caagcccacg
     4981 gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc
     5041 gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc
     5101 gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg
     5161 gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg
     5221 ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa
     5281 cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc
     5341 gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga
     5401 gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac
     5461 ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga
     5521 ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgaa atcaacctct ggattacaaa
     5581 atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac
     5641 gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc
     5701 ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt
     5761 ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc
     5821 tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc
     5881 gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg
     5941 gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg cctgtgttgc cacctggatt
     6001 ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc
     6061 cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt
     6121 cggatctccc tttgggccgc ctccccgcct ggtaccttta agaccaatga cttacaaggc
     6181 agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc
     6241 ccaacgaaga taagatctgc tttttgcttg tactgggtct ctctggttag accagatctg
     6301 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
     6361 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
     6421 cagacccttt tagtcagtgt ggaaaatctc tagcagtagt agttcatgtc atcttattat
     6481 tcagtattta taacttgcaa agaaatgaat atcagagagt gagaggaact tgtttattgc
     6541 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     6601 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctggct
     6661 ctagctatcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     6721 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     6781 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctagactt ttgcagagac
     6841 caaattcgta atcatgtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     6901 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     6961 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     7021 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     7081 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     7141 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     7201 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     7261 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     7321 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     7381 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     7441 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     7501 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     7561 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     7621 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     7681 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     7741 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     7801 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     7861 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     7921 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     7981 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     8041 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     8101 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     8161 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     8221 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     8281 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     8341 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     8401 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     8461 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     8521 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     8581 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     8641 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     8701 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     8761 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     8821 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     8881 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     8941 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     9001 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     9061 cacgaggccc tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     9121 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     9181 gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg catcagagca
     9241 gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg taaggagaaa
     9301 ataccgcatc aggcgccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt
     9361 gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag
     9421 ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgccaagct
     9481 g
//
