LOCUS       V024825                 9327 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V024825
VERSION     V024825
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    1798..1915
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
                     /label="cPPT/CTS"
     promoter        2005..2208
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     regulatory      2279..2288
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             2288..2359
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags"
                     /transl_table=1
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
                     /label="3xFLAG"
     CDS             2366..3442
                     /label="P2RY8(NM_178129)"
                     /note="P2RY8(NM_178129)"
                     /gene="P2RY8"
     promoter        3486..3697
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3710..3978
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      4007..4016
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             4037..4699
                     /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             4769..4822
                     /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             4823..5422
                     /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    5423..6010
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6084..6317
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6389..6510
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6529..6664
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6702..6718)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6726..6742
                     /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(6750..6780)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6795..6816
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7104..7692)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7863..8723)
                     /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(8724..8828)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9302..9318
                     /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 ggttaacttt
     1801 taaaagaaaa ggggggattg gggggtacag tgcaggggaa agaatagtag acataatagc
     1861 aacagacata caaactaaag aattacaaaa acaaattaca aaaattcaaa attttatcga
     1921 tactagtatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
     1981 gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga
     2041 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
     2101 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg
     2161 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctcg tttagtgaac
     2221 cgtcagatcg cctggagacg ccatccacgc tgttttgacc tccatagaag attctagagc
     2281 caccatggac tacaaggatg acgatgacaa ggattacaaa gacgacgatg ataaggacta
     2341 taaggatgat gacgacaaag aattcatgca ggtcccgaac agcaccggcc cggacaacgc
     2401 gacgctgcag atgctgcgga acccggcgat cgcggtggcc ctgcccgtgg tgtactcgct
     2461 ggtggcggcg gtcagcatcc cgggcaacct cttctctctg tgggtgctgt gccggcgcat
     2521 ggggcccaga tccccgtcgg tcatcttcat gatcaacctg agcgtcacgg acctgatgct
     2581 ggccagcgtg ttgcctttcc aaatctacta ccattgcaac cgccaccact gggtattcgg
     2641 ggtgctgctt tgcaacgtgg tgaccgtggc cttttacgca aacatgtatt ccagcatcct
     2701 caccatgacc tgtatcagcg tggagcgctt cctgggggtc ctgtacccgc tcagctccaa
     2761 gcgctggcgc cgccgtcgtt acgcggtggc cgcgtgtgca gggacctggc tgctgctcct
     2821 gaccgccctg tccccgctgg cgcgcaccga tctcacctac ccggtgcacg ccctgggcat
     2881 catcacctgc ttcgacgtcc tcaagtggac gatgctcccc agcgtggcca tgtgggccgt
     2941 gttcctcttc accatcttca tcctgctgtt cctcatcccg ttcgtgatca ccgtggcttg
     3001 ttacacggcc accatcctca agctgttgcg cacggaggag gcgcacggcc gggagcagcg
     3061 gaggcgcgcg gtgggcctgg ccgcggtggt cttgctggcc tttgtcacct gcttcgcccc
     3121 caacaacttc gtgctcctgg cgcacatcgt gagccgcctg ttctacggca agagctacta
     3181 ccacgtgtac aagctcacgc tgtgtctcag ctgcctcaac aactgtctgg acccgtttgt
     3241 ttattacttt gcgtcccggg aattccagct gcgcctgcgg gaatatttgg gctgccgccg
     3301 ggtgcccaga gacaccctgg acacgcgccg cgagagcctc ttctccgcca ggaccacgtc
     3361 cgtgcgctcc gaggccggtg cgcaccctga agggatggag ggagccacca ggcccggcct
     3421 ccagaggcag gagagtgtgt tctgagcggc cgcaaggatc tgcgatcgct ccggtgcccg
     3481 tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa
     3541 ttgaacgggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg
     3601 gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa
     3661 cgttcttttt cgcaacgggt ttgccgccag aacacagctg aagcttcgag gggctcgcat
     3721 ctctccttca cgcgcccgcc gccctacctg aggccgccat ccacgccggt tgagtcgcgt
     3781 tctgccgcct cccgcctgtg gtgcctcctg aactgcgtcc gccgtctagg taagtttaaa
     3841 gctcaggtcg agaccgggcc tttgtccggc gctcccttgg agcctaccta gactcagccg
     3901 gctctccacg ctttgcctga ccctgcttgc tcaactctac gtctttgttt cgttttctgt
     3961 tctgcgccgt tacagatcca agctgtgacc ggcgcctacg ctagacgcca ccatggagag
     4021 cgacgagagc ggcctgcccg ccatggagat cgagtgccgc atcaccggca ccctgaacgg
     4081 cgtggagttc gagctggtgg gcggcggaga gggcaccccc aagcagggcc gcatgaccaa
     4141 caagatgaag agcaccaaag gcgccctgac cttcagcccc tacctgctga gccacgtgat
     4201 gggctacggc ttctaccact tcggcaccta ccccagcggc tacgagaacc ccttcctgca
     4261 cgccatcaac aacggcggct acaccaacac ccgcatcgag aagtacgagg acggcggcgt
     4321 gctgcacgtg agcttcagct accgctacga ggccggccgc gtgatcggcg acttcaaggt
     4381 ggtgggcacc ggcttccccg aggacagcgt gatcttcacc gacaagatca tccgcagcaa
     4441 cgccaccgtg gagcacctgc accccatggg cgataacgtg ctggtgggca gcttcgcccg
     4501 caccttcagc ctgcgcgacg gcggctacta cagcttcgtg gtggacagcc acatgcactt
     4561 caagagcgcc atccacccca gcatcctgca gaacgggggc cccatgttcg ccttccgccg
     4621 cgtggaggag ctgcacagca acaccgagct gggcatcgtg gagtaccagc acgccttcaa
     4681 gacccccatc gccttcgcca gatcccgcgc tcagtcgtcc aattctgccg tggacggcac
     4741 cgccggaccc ggctccaccg gatctcgcga gggcagagga agtcttctaa catgcggtga
     4801 cgtggaggag aatcccggcc ctatgaccga gtacaagccc acggtgcgcc tcgccacccg
     4861 cgacgacgtc cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac
     4921 gcgccacacc gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt
     4981 cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt
     5041 ggcggtctgg accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc
     5101 gcgcatggcc gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct
     5161 ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga
     5221 ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg
     5281 cgccggggtg cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg
     5341 gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat
     5401 gacccgcaag cccggtgcct gaaatcaacc tctggattac aaaatttgtg aaagattgac
     5461 tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt
     5521 gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt
     5581 gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt
     5641 gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg
     5701 gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg
     5761 ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaagct
     5821 gacgtccttt ccatggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt
     5881 ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc
     5941 tctgcggcct cttccgcgtc tccgccttcg ccctcagacg agtcggatct ccctttggcc
     6001 gcctccccgc ctggtacctt taagaccaat gacttacaag gcagctgtag atcttagcca
     6061 ctttttaaaa gaaaaggggg gactggaagg gctaattcac tcccaacgaa aataagatct
     6121 gctttttgct tgtactgggt ctctctggtt agaccagatc tgagcctggg agctctctgg
     6181 ctaactaggg aacccactgc ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt
     6241 gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc ctcagaccct tttagtcagt
     6301 gtggaaaatc tctagcagta gtagttcatg tcatcttatt attcagtatt tataacttgc
     6361 aaagaaatga atatcagaga gtgagaggaa cttgtttatt gcagcttata atggttacaa
     6421 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     6481 tggtttgtcc aaactcatca atgtatctta tcatgtctgg ctctagctat cccgccccta
     6541 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     6601 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     6661 ggcctagact tttgcagaga cggcccaaat tcgtaatcat ggtcatagct gtttcctgtg
     6721 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     6781 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     6841 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     6901 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     6961 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     7021 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     7081 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     7141 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     7201 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     7261 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     7321 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     7381 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     7441 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     7501 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
     7561 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     7621 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     7681 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     7741 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     7801 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     7861 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     7921 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     7981 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     8041 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     8101 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     8161 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     8221 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     8281 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     8341 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     8401 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     8461 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     8521 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     8581 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     8641 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     8701 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     8761 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     8821 gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg
     8881 acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat
     8941 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     9001 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     9061 tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa
     9121 ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc attcaggctg
     9181 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa
     9241 gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt
     9301 tgtaaaacga cggccagtgc caagctg
//
