LOCUS       V042250                 8692 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V042250
VERSION     V042250
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..2718
                     /label="ID2(NM_002166)"
                     /note="ID2(NM_002166)"
                     /gene="ID2"
     CDS             2725..2790
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        2834..3045
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3058..3326
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      3355..3364
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             3385..4047
                     /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             4117..4170
                     /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             4171..4770
                     /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    4771..5359
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5433..5666
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    5738..5859
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      5899..6034
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6067..6083)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6091..6107
                     /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(6115..6145)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6160..6181
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6469..7057)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7228..8088)
                     /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(8089..8193)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     8667..8683
                     /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 aagccttcag tcccgtgagg
     2341 tccgttagga aaaacagcct gtcggaccac agcctgggca tctcccggag caaaacccct
     2401 gtggacgacc cgatgagcct gctatacaac atgaacgact gctactccaa gctcaaggag
     2461 ctggtgccca gcatccccca gaacaagaag gtgagcaaga tggaaatcct gcagcacgtc
     2521 atcgactaca tcttggacct gcagatcgcc ctggactcgc atcccactat tgtcagcctg
     2581 catcaccaga gacccgggca gaaccaggcg tccaggacgc cgctgaccac cctcaacacg
     2641 gatatcagca tcctgtcctt gcaggcttct gaattccctt ctgagttaat gtcaaatgac
     2701 agcaaagcac tgtgtggcgg atccgactac aaagaccatg acggtgatta taaagatcat
     2761 gacatcgact acaaggatga cgatgacaag tagcggccgc gaaggatctg cgatcgctcc
     2821 ggtgcccgtc agtgggcaga gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg
     2881 gtcggcaatt gaacgggtgc ctagagaagg tggcgcgggg taaactggga aagtgatgtc
     2941 gtgtactggc tccgcctttt tcccgagggt gggggagaac cgtatataag tgcagtagtc
     3001 gccgtgaacg ttctttttcg caacgggttt gccgccagaa cacagctgaa gcttcgaggg
     3061 gctcgcatct ctccttcacg cgcccgccgc cctacctgag gccgccatcc acgccggttg
     3121 agtcgcgttc tgccgcctcc cgcctgtggt gcctcctgaa ctgcgtccgc cgtctaggta
     3181 agtttaaagc tcaggtcgag accgggcctt tgtccggcgc tcccttggag cctacctaga
     3241 ctcagccggc tctccacgct ttgcctgacc ctgcttgctc aactctacgt ctttgtttcg
     3301 ttttctgttc tgcgccgtta cagatccaag ctgtgaccgg cgcctacgct agacgccacc
     3361 atggagagcg acgagagcgg cctgcccgcc atggagatcg agtgccgcat caccggcacc
     3421 ctgaacggcg tggagttcga gctggtgggc ggcggagagg gcacccccaa gcagggccgc
     3481 atgaccaaca agatgaagag caccaaaggc gccctgacct tcagccccta cctgctgagc
     3541 cacgtgatgg gctacggctt ctaccacttc ggcacctacc ccagcggcta cgagaacccc
     3601 ttcctgcacg ccatcaacaa cggcggctac accaacaccc gcatcgagaa gtacgaggac
     3661 ggcggcgtgc tgcacgtgag cttcagctac cgctacgagg ccggccgcgt gatcggcgac
     3721 ttcaaggtgg tgggcaccgg cttccccgag gacagcgtga tcttcaccga caagatcatc
     3781 cgcagcaacg ccaccgtgga gcacctgcac cccatgggcg ataacgtgct ggtgggcagc
     3841 ttcgcccgca ccttcagcct gcgcgacggc ggctactaca gcttcgtggt ggacagccac
     3901 atgcacttca agagcgccat ccaccccagc atcctgcaga acgggggccc catgttcgcc
     3961 ttccgccgcg tggaggagct gcacagcaac accgagctgg gcatcgtgga gtaccagcac
     4021 gccttcaaga cccccatcgc cttcgccaga tcccgcgctc agtcgtccaa ttctgccgtg
     4081 gacggcaccg ccggacccgg ctccaccgga tctcgcgagg gcagaggaag tcttctaaca
     4141 tgcggtgacg tggaggagaa tcccggccct atgaccgagt acaagcccac ggtgcgcctc
     4201 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac
     4261 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa
     4321 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc
     4381 gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag
     4441 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa
     4501 ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc
     4561 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg
     4621 gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc
     4681 tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc
     4741 tggtgcatga cccgcaagcc cggtgcctga aatcaacctc tggattacaa aatttgtgaa
     4801 agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta
     4861 atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa
     4921 tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg
     4981 tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc
     5041 ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc
     5101 cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg
     5161 gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg
     5221 acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg
     5281 ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc
     5341 ctttgggccg cctccccgcc tggtaccttt aagaccaatg acttacaagg cagctgtaga
     5401 tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag
     5461 ataagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga
     5521 gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct
     5581 tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt
     5641 ttagtcagtg tggaaaatct ctagcagtag tagttcatgt catcttatta ttcagtattt
     5701 ataacttgca aagaaatgaa tatcagagag tgagaggaac ttgtttattg cagcttataa
     5761 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     5821 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctggc tctagctatc
     5881 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     5941 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     6001 ttccagaagt agtgaggagg cttttttgga ggcctagact tttgcagaga ccaaattcgt
     6061 aatcatgtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     6121 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     6181 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     6241 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     6301 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     6361 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     6421 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     6481 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     6541 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     6601 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     6661 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     6721 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     6781 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     6841 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     6901 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     6961 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     7021 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     7081 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     7141 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     7201 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     7261 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     7321 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     7381 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     7441 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     7501 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     7561 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     7621 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     7681 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     7741 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     7801 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     7861 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     7921 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     7981 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     8041 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     8101 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     8161 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     8221 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
     8281 ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
     8341 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
     8401 agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact
     8461 gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     8521 caggcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
     8581 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
     8641 gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgccaagc tg
//
