LOCUS       V035250                 8902 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V035250
VERSION     V035250
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..2928
                     /label="Crk(NM_001277219)"
                     /note="Crk(NM_001277219)"
                     /gene="Crk"
     CDS             2935..3000
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3044..3255
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3268..3536
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      3565..3574
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             3595..4257
                     /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             4327..4380
                     /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             4381..4980
                     /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    4981..5569
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5643..5876
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    5948..6069
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6109..6244
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6277..6293)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6301..6317
                     /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(6325..6355)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6370..6391
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6679..7267)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7438..8298)
                     /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(8299..8403)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     8877..8893
                     /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 gccaccatgg cgggcaactt cgactcggag
     2341 gagcggagta gctggtactg gggccgcctg agccggcagg aggcggtggc gctattgcag
     2401 ggccagcggc acggggtgtt cctggtgcgg gactcgagca ccagccccgg ggactatgtg
     2461 cttagcgtct ccgaaaactc gcgcgtctcc cactacatca tcaacagcag cggcccgcgc
     2521 cctccagtgc ctccgtcgcc cgctcagcct ccgccgggag tgagtccctc caggctccga
     2581 ataggagatc aagaatttga ttcattgcct gctttactgg aattctacaa aatacactat
     2641 ttggacacta caacattgat agaaccagtg gccagatcaa ggcagggtag tggagtgatt
     2701 ctcaggcagg aggaggcaga gtatgtgcgg gccctctttg actttaatgg gaatgatgaa
     2761 gaagatcttc cctttaagaa aggagacatc ctgagaatcc gggataagcc tgaagagcag
     2821 tggtggaatg cagaggacag cgaaggaaag agggggatga ttcctgtccc ttacgtggag
     2881 aagtatagac ctgcctccgc ctcagtatcg gctctgattg gaggtcgggg atccgactac
     2941 aaagaccatg acggtgatta taaagatcat gacatcgact acaaggatga cgatgacaag
     3001 tagcggccgc gaaggatctg cgatcgctcc ggtgcccgtc agtgggcaga gcgcacatcg
     3061 cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt gaacgggtgc ctagagaagg
     3121 tggcgcgggg taaactggga aagtgatgtc gtgtactggc tccgcctttt tcccgagggt
     3181 gggggagaac cgtatataag tgcagtagtc gccgtgaacg ttctttttcg caacgggttt
     3241 gccgccagaa cacagctgaa gcttcgaggg gctcgcatct ctccttcacg cgcccgccgc
     3301 cctacctgag gccgccatcc acgccggttg agtcgcgttc tgccgcctcc cgcctgtggt
     3361 gcctcctgaa ctgcgtccgc cgtctaggta agtttaaagc tcaggtcgag accgggcctt
     3421 tgtccggcgc tcccttggag cctacctaga ctcagccggc tctccacgct ttgcctgacc
     3481 ctgcttgctc aactctacgt ctttgtttcg ttttctgttc tgcgccgtta cagatccaag
     3541 ctgtgaccgg cgcctacgct agacgccacc atggagagcg acgagagcgg cctgcccgcc
     3601 atggagatcg agtgccgcat caccggcacc ctgaacggcg tggagttcga gctggtgggc
     3661 ggcggagagg gcacccccaa gcagggccgc atgaccaaca agatgaagag caccaaaggc
     3721 gccctgacct tcagccccta cctgctgagc cacgtgatgg gctacggctt ctaccacttc
     3781 ggcacctacc ccagcggcta cgagaacccc ttcctgcacg ccatcaacaa cggcggctac
     3841 accaacaccc gcatcgagaa gtacgaggac ggcggcgtgc tgcacgtgag cttcagctac
     3901 cgctacgagg ccggccgcgt gatcggcgac ttcaaggtgg tgggcaccgg cttccccgag
     3961 gacagcgtga tcttcaccga caagatcatc cgcagcaacg ccaccgtgga gcacctgcac
     4021 cccatgggcg ataacgtgct ggtgggcagc ttcgcccgca ccttcagcct gcgcgacggc
     4081 ggctactaca gcttcgtggt ggacagccac atgcacttca agagcgccat ccaccccagc
     4141 atcctgcaga acgggggccc catgttcgcc ttccgccgcg tggaggagct gcacagcaac
     4201 accgagctgg gcatcgtgga gtaccagcac gccttcaaga cccccatcgc cttcgccaga
     4261 tcccgcgctc agtcgtccaa ttctgccgtg gacggcaccg ccggacccgg ctccaccgga
     4321 tctcgcgagg gcagaggaag tcttctaaca tgcggtgacg tggaggagaa tcccggccct
     4381 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta
     4441 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac
     4501 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     4561 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     4621 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     4681 tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag
     4741 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc
     4801 agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg
     4861 gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     4921 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga
     4981 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
     5041 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
     5101 atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg
     5161 tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact
     5221 ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct
     5281 attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg
     5341 ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc
     5401 gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc
     5461 aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt
     5521 cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcc tggtaccttt
     5581 aagaccaatg acttacaagg cagctgtaga tcttagccac tttttaaaag aaaagggggg
     5641 actggaaggg ctaattcact cccaacgaag ataagatctg ctttttgctt gtactgggtc
     5701 tctctggtta gaccagatct gagcctggga gctctctggc taactaggga acccactgct
     5761 taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga
     5821 ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagtag
     5881 tagttcatgt catcttatta ttcagtattt ataacttgca aagaaatgaa tatcagagag
     5941 tgagaggaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     6001 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     6061 tgtatcttat catgtctggc tctagctatc ccgcccctaa ctccgcccat cccgccccta
     6121 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     6181 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     6241 ggcctagact tttgcagaga ccaaattcgt aatcatgtca tagctgtttc ctgtgtgaaa
     6301 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     6361 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     6421 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     6481 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     6541 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     6601 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     6661 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     6721 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     6781 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     6841 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     6901 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     6961 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     7021 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     7081 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     7141 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     7201 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     7261 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     7321 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     7381 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     7441 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     7501 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     7561 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     7621 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     7681 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     7741 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     7801 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     7861 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     7921 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     7981 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     8041 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     8101 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     8161 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     8221 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     8281 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     8341 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     8401 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
     8461 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg
     8521 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     8581 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
     8641 taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc
     8701 gcacagatgc gtaaggagaa aataccgcat caggcgccat tcgccattca ggctgcgcaa
     8761 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg
     8821 atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa
     8881 aacgacggcc agtgccaagc tg
//
