LOCUS       V032069                 9475 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V032069
VERSION     V032069
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..3501
                     /label="DHODH(NM_001361)"
                     /note="DHODH(NM_001361)"
                     /gene="DHODH"
     CDS             3508..3573
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3617..3828
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3841..4109
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      4138..4147
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             4168..4830
                     /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             4900..4953
                     /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             4954..5553
                     /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    5554..6142
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6216..6449
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6521..6642
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6682..6817
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6850..6866)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6874..6890
                     /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(6898..6928)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6943..6964
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7252..7840)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8011..8871)
                     /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(8872..8976)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9450..9466
                     /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 cgtggagaca cctgaaaaag
     2341 cgggcccagg atgctgtgat catcctgggg ggaggaggac ttctcttcgc ctcctacctg
     2401 atggccacgg gagatgagcg tttctatgct gaacacctga tgccgactct gcaggggctg
     2461 ctggacccgg agtcagccca cagactggct gttcgcttca cctccctggg gctccttcca
     2521 cgggccagat ttcaagactc tgacatgctg gaagtgagag ttctgggcca taaattccga
     2581 aatccagtag gaattgctgc aggatttgac aagcatgggg aagccgtgga cggactttat
     2641 aagatgggct ttggttttgt tgagatagga agtgtgactc caaaacctca ggaaggaaac
     2701 cctagaccca gagtcttccg cctccctgag gaccaagctg tcattaacag gtatggattt
     2761 aacagtcacg ggctttcagt ggtggaacac aggttacggg ccagacagca gaagcaggcc
     2821 aagctcacag aagatggact gcctctgggg gtcaacttgg ggaagaacaa gacctcagtg
     2881 gacgccgcgg aggactacgc agaaggggtg cgcgtactgg gccccctggc cgactacctg
     2941 gtggtgaatg tgtccagccc caacactgcc gggctgcgga gccttcaggg aaaggccgag
     3001 ctgcgccgcc tgctgaccaa ggtgctgcag gagagggatg gcttgcggag agtgcacagg
     3061 ccggcagtcc tggtgaagat cgctcctgac ctcaccagcc aggataagga ggacattgcc
     3121 agtgtggtca aagagttggg catcgatggg ctgattgtta cgaacaccac cgtgagtcgc
     3181 cctgcgggcc tccagggtgc cctgcgctct gaaacaggag ggctgagtgg gaagcccctc
     3241 cgggatttat caactcaaac cattcgggag atgtatgcac tcacccaagg ccgagttccc
     3301 ataattgggg ttggtggtgt gagcagcggg caggacgcgc tggagaagat ccgggcaggg
     3361 gcctccctgg tgcagctgta cacggccctc accttctggg ggccacccgt tgtgggcaaa
     3421 gtcaagcggg aactggaggc ccttctgaaa gagcagggct ttggcggagt cacagatgcc
     3481 attggagcag atcatcggag gggatccgac tacaaagacc atgacggtga ttataaagat
     3541 catgacatcg actacaagga tgacgatgac aagtagcggc cgcgaaggat ctgcgatcgc
     3601 tccggtgccc gtcagtgggc agagcgcaca tcgcccacag tccccgagaa gttgggggga
     3661 ggggtcggca attgaacggg tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat
     3721 gtcgtgtact ggctccgcct ttttcccgag ggtgggggag aaccgtatat aagtgcagta
     3781 gtcgccgtga acgttctttt tcgcaacggg tttgccgcca gaacacagct gaagcttcga
     3841 ggggctcgca tctctccttc acgcgcccgc cgccctacct gaggccgcca tccacgccgg
     3901 ttgagtcgcg ttctgccgcc tcccgcctgt ggtgcctcct gaactgcgtc cgccgtctag
     3961 gtaagtttaa agctcaggtc gagaccgggc ctttgtccgg cgctcccttg gagcctacct
     4021 agactcagcc ggctctccac gctttgcctg accctgcttg ctcaactcta cgtctttgtt
     4081 tcgttttctg ttctgcgccg ttacagatcc aagctgtgac cggcgcctac gctagacgcc
     4141 accatggaga gcgacgagag cggcctgccc gccatggaga tcgagtgccg catcaccggc
     4201 accctgaacg gcgtggagtt cgagctggtg ggcggcggag agggcacccc caagcagggc
     4261 cgcatgacca acaagatgaa gagcaccaaa ggcgccctga ccttcagccc ctacctgctg
     4321 agccacgtga tgggctacgg cttctaccac ttcggcacct accccagcgg ctacgagaac
     4381 cccttcctgc acgccatcaa caacggcggc tacaccaaca cccgcatcga gaagtacgag
     4441 gacggcggcg tgctgcacgt gagcttcagc taccgctacg aggccggccg cgtgatcggc
     4501 gacttcaagg tggtgggcac cggcttcccc gaggacagcg tgatcttcac cgacaagatc
     4561 atccgcagca acgccaccgt ggagcacctg caccccatgg gcgataacgt gctggtgggc
     4621 agcttcgccc gcaccttcag cctgcgcgac ggcggctact acagcttcgt ggtggacagc
     4681 cacatgcact tcaagagcgc catccacccc agcatcctgc agaacggggg ccccatgttc
     4741 gccttccgcc gcgtggagga gctgcacagc aacaccgagc tgggcatcgt ggagtaccag
     4801 cacgccttca agacccccat cgccttcgcc agatcccgcg ctcagtcgtc caattctgcc
     4861 gtggacggca ccgccggacc cggctccacc ggatctcgcg agggcagagg aagtcttcta
     4921 acatgcggtg acgtggagga gaatcccggc cctatgaccg agtacaagcc cacggtgcgc
     4981 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac
     5041 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg
     5101 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac
     5161 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc
     5221 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg
     5281 gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc
     5341 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
     5401 gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc
     5461 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
     5521 acctggtgca tgacccgcaa gcccggtgcc tgaaatcaac ctctggatta caaaatttgt
     5581 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     5641 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     5701 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     5761 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     5821 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     5881 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     5941 tcggggaaat catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc
     6001 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     6061 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     6121 tccctttggg ccgcctcccc gcctggtacc tttaagacca atgacttaca aggcagctgt
     6181 agatcttagc cactttttaa aagaaaaggg gggactggaa gggctaattc actcccaacg
     6241 aagataagat ctgctttttg cttgtactgg gtctctctgg ttagaccaga tctgagcctg
     6301 ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt
     6361 gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc
     6421 cttttagtca gtgtggaaaa tctctagcag tagtagttca tgtcatctta ttattcagta
     6481 tttataactt gcaaagaaat gaatatcaga gagtgagagg aacttgttta ttgcagctta
     6541 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     6601 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggctctagct
     6661 atcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     6721 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
     6781 ctattccaga agtagtgagg aggctttttt ggaggcctag acttttgcag agaccaaatt
     6841 cgtaatcatg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     6901 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     6961 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     7021 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     7081 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     7141 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     7201 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     7261 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     7321 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     7381 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     7441 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     7501 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     7561 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     7621 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     7681 ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     7741 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     7801 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     7861 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     7921 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     7981 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     8041 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     8101 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     8161 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     8221 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     8281 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     8341 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     8401 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     8461 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     8521 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     8581 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     8641 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     8701 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     8761 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     8821 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     8881 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     8941 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     9001 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
     9061 gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
     9121 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
     9181 gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt
     9241 actgagagtg caccatatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg
     9301 catcaggcgc cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc
     9361 ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt
     9421 aacgccaggg ttttcccagt cacgacgttg taaaacgacg gccagtgcca agctg
//
