LOCUS       V035566                 9181 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V035566
VERSION     V035566
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..3207
                     /label="Cdk1(NM_007659)"
                     /note="Cdk1(NM_007659)"
                     /gene="Cdk1"
     CDS             3214..3279
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3323..3534
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3547..3815
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      3844..3853
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             3874..4536
                     /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             4606..4659
                     /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             4660..5259
                     /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    5260..5848
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5922..6155
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6227..6348
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6388..6523
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6556..6572)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6580..6596
                     /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(6604..6634)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6649..6670
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6958..7546)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7717..8577)
                     /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(8578..8682)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9156..9172
                     /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 aagactatat caaaatagag
     2341 aaaattggag aaggtactta cggtgtggtg tataagggta gacacagagt cactggccag
     2401 atagtggcca tgaagaagat cagacttgaa agcgaggaag aaggagtgcc cagtactgca
     2461 attcgggaaa tctctctatt aaaagaactt cgacatccaa atatagtcag cctgcaggat
     2521 gtgctcatgc aggactccag gctgtatctc atctttgagt tcctgtccat ggacctcaag
     2581 aagtacctgg actccatccc tcctgggcag ttcatggatt cttcactcgt taagagttac
     2641 ttacaccaaa tcctccaggg aattgtgttt tgccactccc ggcgagttct tcacagagac
     2701 ttgaaacctc aaaatctatt gattgatgac aaaggaacaa tcaaactggc tgatttcggc
     2761 cttgccagag cgtttggaat accgatacga gtgtacacac acgaggtagt gacgctgtgg
     2821 taccgatctc cagaagtgtt gctgggctcg gctcgttact ccactccggt tgacatctgg
     2881 agtataggga ccatatttgc agaactggcc accaagaagc cgcttttcca cggcgactca
     2941 gagattgacc agctcttcag gatcttcaga gctctgggca ctcctaacaa cgaagtgtgg
     3001 ccagaagtcg agtccctgca ggactacaag aacacctttc ccaagtggaa gccggggagc
     3061 ctcgcatccc acgtcaagaa cctggacgag aacggcttgg atttgctctc aaaaatgcta
     3121 gtctatgatc ctgccaaacg aatctctggc aaaatggccc tgaagcaccc gtactttgat
     3181 gacttggaca atcagattaa gaagatggga tccgactaca aagaccatga cggtgattat
     3241 aaagatcatg acatcgacta caaggatgac gatgacaagt agcggccgcg aaggatctgc
     3301 gatcgctccg gtgcccgtca gtgggcagag cgcacatcgc ccacagtccc cgagaagttg
     3361 gggggagggg tcggcaattg aacgggtgcc tagagaaggt ggcgcggggt aaactgggaa
     3421 agtgatgtcg tgtactggct ccgccttttt cccgagggtg ggggagaacc gtatataagt
     3481 gcagtagtcg ccgtgaacgt tctttttcgc aacgggtttg ccgccagaac acagctgaag
     3541 cttcgagggg ctcgcatctc tccttcacgc gcccgccgcc ctacctgagg ccgccatcca
     3601 cgccggttga gtcgcgttct gccgcctccc gcctgtggtg cctcctgaac tgcgtccgcc
     3661 gtctaggtaa gtttaaagct caggtcgaga ccgggccttt gtccggcgct cccttggagc
     3721 ctacctagac tcagccggct ctccacgctt tgcctgaccc tgcttgctca actctacgtc
     3781 tttgtttcgt tttctgttct gcgccgttac agatccaagc tgtgaccggc gcctacgcta
     3841 gacgccacca tggagagcga cgagagcggc ctgcccgcca tggagatcga gtgccgcatc
     3901 accggcaccc tgaacggcgt ggagttcgag ctggtgggcg gcggagaggg cacccccaag
     3961 cagggccgca tgaccaacaa gatgaagagc accaaaggcg ccctgacctt cagcccctac
     4021 ctgctgagcc acgtgatggg ctacggcttc taccacttcg gcacctaccc cagcggctac
     4081 gagaacccct tcctgcacgc catcaacaac ggcggctaca ccaacacccg catcgagaag
     4141 tacgaggacg gcggcgtgct gcacgtgagc ttcagctacc gctacgaggc cggccgcgtg
     4201 atcggcgact tcaaggtggt gggcaccggc ttccccgagg acagcgtgat cttcaccgac
     4261 aagatcatcc gcagcaacgc caccgtggag cacctgcacc ccatgggcga taacgtgctg
     4321 gtgggcagct tcgcccgcac cttcagcctg cgcgacggcg gctactacag cttcgtggtg
     4381 gacagccaca tgcacttcaa gagcgccatc caccccagca tcctgcagaa cgggggcccc
     4441 atgttcgcct tccgccgcgt ggaggagctg cacagcaaca ccgagctggg catcgtggag
     4501 taccagcacg ccttcaagac ccccatcgcc ttcgccagat cccgcgctca gtcgtccaat
     4561 tctgccgtgg acggcaccgc cggacccggc tccaccggat ctcgcgaggg cagaggaagt
     4621 cttctaacat gcggtgacgt ggaggagaat cccggcccta tgaccgagta caagcccacg
     4681 gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc
     4741 gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc
     4801 gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg
     4861 gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg
     4921 ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa
     4981 cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc
     5041 gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga
     5101 gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac
     5161 ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga
     5221 ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgaa atcaacctct ggattacaaa
     5281 atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac
     5341 gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc
     5401 ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt
     5461 ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc
     5521 tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc
     5581 gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg
     5641 gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg cctgtgttgc cacctggatt
     5701 ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc
     5761 cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt
     5821 cggatctccc tttgggccgc ctccccgcct ggtaccttta agaccaatga cttacaaggc
     5881 agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc
     5941 ccaacgaaga taagatctgc tttttgcttg tactgggtct ctctggttag accagatctg
     6001 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
     6061 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
     6121 cagacccttt tagtcagtgt ggaaaatctc tagcagtagt agttcatgtc atcttattat
     6181 tcagtattta taacttgcaa agaaatgaat atcagagagt gagaggaact tgtttattgc
     6241 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     6301 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctggct
     6361 ctagctatcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     6421 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     6481 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctagactt ttgcagagac
     6541 caaattcgta atcatgtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     6601 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     6661 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     6721 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     6781 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     6841 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     6901 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     6961 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     7021 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     7081 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     7141 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     7201 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     7261 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     7321 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     7381 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     7441 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     7501 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     7561 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     7621 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     7681 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     7741 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     7801 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     7861 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     7921 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     7981 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     8041 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     8101 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     8161 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     8221 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     8281 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     8341 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     8401 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     8461 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     8521 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     8581 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     8641 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     8701 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     8761 cacgaggccc tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     8821 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     8881 gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg catcagagca
     8941 gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg taaggagaaa
     9001 ataccgcatc aggcgccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt
     9061 gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag
     9121 ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgccaagct
     9181 g
//
