LOCUS       V025005                 9521 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V025005
VERSION     V025005
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..3564
                     /label="MKRN2(NM_014160)"
                     /note="MKRN2(NM_014160)"
                     /gene="MKRN2"
     CDS             3571..3636
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3680..3891
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3904..4172
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      4201..4210
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             4231..4893
                     /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             4963..5016
                     /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             5017..5616
                     /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    5617..6204
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6278..6511
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6583..6704
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6723..6858
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6896..6912)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6920..6936
                     /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(6944..6974)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6989..7010
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7298..7886)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8057..8917)
                     /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(8918..9022)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9496..9512
                     /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 gcaccaagca gatcacttgc
     2341 aggtatttta tgcatggtgt gtgtcgggaa ggaagtcagt gcctattctc acatgacttg
     2401 gcaaacagca aaccgtccac catctgcaag tactaccaga agggctactg tgcctatgga
     2461 actcggtgca gatatgacca cacgaggccc tctgctgcag ctggaggtgc tgtgggcacc
     2521 atggcccaca gtgtgccctc cccagctttc cacagtcctc accctccttc cgaggtcact
     2581 gcatccattg tgaaaactaa ctcacatgaa cccggaaagc gtgaaaagag aacattggtt
     2641 cttagagacc gaaatctctc tggcatggct gaaaggaaga cccagccgag catggtgagt
     2701 aatccaggca gctgcagcga cccccagccc agccccgaga tgaagccgca ttcctacctg
     2761 gatgccatca ggagtggcct tgatgacgtg gaggccagca gctcctacag caacgagcag
     2821 cagctgtgcc cctacgcagc tgctggggag tgccggtttg gggatgcctg tgtctacctg
     2881 cacggggagg tgtgtgaaat ctgtaggctg caagtcttgc acccattcga cccagagcag
     2941 aggaaggctc atgaaaagat ctgcatgttg acgttcgaac acgagatgga aaaggccttt
     3001 gccttccagg caagccagga caaagtgtgc agtatctgca tggaagtgat cctggagaag
     3061 gcctctgctt ctgagaggag atttgggatt ctctccaatt gcaatcacac gtactgtttg
     3121 tcctgcatcc ggcagtggcg gtgtgccaaa cagtttgaaa acccaatcat taagtcttgt
     3181 ccagaatgcc gtgtgatatc agagtttgta attccaagtg tgtattgggt ggaagatcag
     3241 aataaaaaga acgagttgat tgaagctttc aaacagggga tggggaaaaa agcctgtaaa
     3301 tactttgagc aaggcaaggg gacctgccca tttggaagca aatgtcttta tcgccatgct
     3361 taccccgatg ggcggctagc agagcctgag aaacctcgga aacagctcag ttctcaaggc
     3421 actgtgaggt tctttaattc agtgcggctc tgggatttca tcgagaaccg agaaagccgg
     3481 catgtcccca acaatgaaga tgtcgacatg acagagctcg gggacctctt catgcacctt
     3541 tctggagtgg aatcatcaga acccggatcc gactacaaag accatgacgg tgattataaa
     3601 gatcatgaca tcgactacaa ggatgacgat gacaagtagc ggccgcgaag gatctgcgat
     3661 cgctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
     3721 ggaggggtcg gcaattgaac gggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
     3781 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
     3841 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca gctgaagctt
     3901 cgaggggctc gcatctctcc ttcacgcgcc cgccgcccta cctgaggccg ccatccacgc
     3961 cggttgagtc gcgttctgcc gcctcccgcc tgtggtgcct cctgaactgc gtccgccgtc
     4021 taggtaagtt taaagctcag gtcgagaccg ggcctttgtc cggcgctccc ttggagccta
     4081 cctagactca gccggctctc cacgctttgc ctgaccctgc ttgctcaact ctacgtcttt
     4141 gtttcgtttt ctgttctgcg ccgttacaga tccaagctgt gaccggcgcc tacgctagac
     4201 gccaccatgg agagcgacga gagcggcctg cccgccatgg agatcgagtg ccgcatcacc
     4261 ggcaccctga acggcgtgga gttcgagctg gtgggcggcg gagagggcac ccccaagcag
     4321 ggccgcatga ccaacaagat gaagagcacc aaaggcgccc tgaccttcag cccctacctg
     4381 ctgagccacg tgatgggcta cggcttctac cacttcggca cctaccccag cggctacgag
     4441 aaccccttcc tgcacgccat caacaacggc ggctacacca acacccgcat cgagaagtac
     4501 gaggacggcg gcgtgctgca cgtgagcttc agctaccgct acgaggccgg ccgcgtgatc
     4561 ggcgacttca aggtggtggg caccggcttc cccgaggaca gcgtgatctt caccgacaag
     4621 atcatccgca gcaacgccac cgtggagcac ctgcacccca tgggcgataa cgtgctggtg
     4681 ggcagcttcg cccgcacctt cagcctgcgc gacggcggct actacagctt cgtggtggac
     4741 agccacatgc acttcaagag cgccatccac cccagcatcc tgcagaacgg gggccccatg
     4801 ttcgccttcc gccgcgtgga ggagctgcac agcaacaccg agctgggcat cgtggagtac
     4861 cagcacgcct tcaagacccc catcgccttc gccagatccc gcgctcagtc gtccaattct
     4921 gccgtggacg gcaccgccgg acccggctcc accggatctc gcgagggcag aggaagtctt
     4981 ctaacatgcg gtgacgtgga ggagaatccc ggccctatga ccgagtacaa gcccacggtg
     5041 cgcctcgcca cccgcgacga cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc
     5101 gactaccccg ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag
     5161 ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac
     5221 gacggcgccg cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc
     5281 gccgagatcg gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag
     5341 atggaaggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc
     5401 ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg
     5461 gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc
     5521 cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg
     5581 cgcacctggt gcatgacccg caagcccggt gcctgaaatc aacctctgga ttacaaaatt
     5641 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
     5701 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
     5761 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc
     5821 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt
     5881 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc
     5941 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg
     6001 ttgtcgggga agctgacgtc ctttccatgg ctgctcgcct gtgttgccac ctggattctg
     6061 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc
     6121 ggcctgctgc cggctctgcg gcctcttccg cgtctccgcc ttcgccctca gacgagtcgg
     6181 atctcccttt ggccgcctcc ccgcctggta cctttaagac caatgactta caaggcagct
     6241 gtagatctta gccacttttt aaaagaaaag gggggactgg aagggctaat tcactcccaa
     6301 cgaaaataag atctgctttt tgcttgtact gggtctctct ggttagacca gatctgagcc
     6361 tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga
     6421 gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga
     6481 cccttttagt cagtgtggaa aatctctagc agtagtagtt catgtcatct tattattcag
     6541 tatttataac ttgcaaagaa atgaatatca gagagtgaga ggaacttgtt tattgcagct
     6601 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     6661 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggctctag
     6721 ctatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     6781 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
     6841 gaggcttttt tggaggccta gacttttgca gagacggccc aaattcgtaa tcatggtcat
     6901 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     6961 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     7021 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     7081 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     7141 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     7201 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     7261 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     7321 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     7381 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     7441 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     7501 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     7561 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     7621 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     7681 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga
     7741 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     7801 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     7861 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     7921 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     7981 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     8041 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     8101 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     8161 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     8221 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     8281 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     8341 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     8401 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     8461 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     8521 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     8581 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     8641 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     8701 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     8761 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     8821 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     8881 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     8941 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     9001 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     9061 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtctcg
     9121 cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag
     9181 cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg
     9241 gcgggtgtcg gggctggctt aactatgcgg catcagagca gattgtactg agagtgcacc
     9301 atatgcggtg tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgccatt
     9361 cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac
     9421 gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt
     9481 cccagtcacg acgttgtaaa acgacggcca gtgccaagct g
//
