LOCUS       V045210                 9880 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V045210
VERSION     V045210
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..1920
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
                     /label="cPPT/CTS"
     enhancer        1933..2236
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        2237..2440
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     CDS             2475..5099
                     /label="KLHL29(NM_052920)"
                     /note="KLHL29(NM_052920)"
                     /gene="KLHL29"
     CDS             5106..5171
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             5181..5237
                     /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 porcine teschovirus-1
                     polyprotein"
                     /transl_table=1
                     /translation="ATNFSLLKQAGDVEENPGP"
                     /label="P2A"
     CDS             5250..5969
                     /codon_start=1
                     /note="mammalian codon-optimized"
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /transl_table=1
                     /translation="M,V,SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKL
                     TLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFK
                     DDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNG
                     IKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMV
                     LLEFVTAAGITLGMDELYK*"
                     /label="EGFP"
     misc_feature    5976..6563
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6637..6870
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6942..7063
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      7082..7217
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(7255..7271)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    7279..7295
                     /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(7303..7333)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    7348..7369
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7657..8245)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8416..9276)
                     /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(9277..9381)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9855..9871
                     /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 attacaaaat tcaaaatttt
     1921 atcgatacta gtcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
     1981 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
     2041 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
     2101 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
     2161 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
     2221 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
     2281 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
     2341 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
     2401 cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat
     2461 ccgctagcgc caccatgtcc cggcaccata gccgcttcga aagagattac cgggtgggct
     2521 gggaccgccg cgaatggagc gtcaacggga cgcatgggac caccagcatc tgcagtgtca
     2581 cctcgggggc cggtggcggc acagccagca gcctcagcgt ccggcccggc ctcctgccgc
     2641 tgcccgtggt gccctcccgg ctgcccaccc cggctacagc tcctgctccc tgcaccaccg
     2701 gcagcagcga ggccatcacc agcctcgtgg ccagctctgc gtctgcggtc accaccaagg
     2761 ctcccggcat ctccaaaggg gacagtcagt cccagggact ggcgaccagc atccggtggg
     2821 ggcagacgcc tatcaatcag tccacaccct gggacactga tgagccaccc tccaaacaga
     2881 tgagagagag tgacaatcca ggcacagggc catgggtgac cacggtggcc gccgggaacc
     2941 agcccaccct gattgcacac tcctatggag tggcccagcc tcccaccttc agcccggctg
     3001 tgaacgtcca ggccccggtc attggggtga ccccctcact gcctccccac gtggggcccc
     3061 agctcccgct gatgccaggc cactactcgc tccctcagcc gccctctcag ccactgagca
     3121 gcgtggtggt caacatgcct gcccaggccc tgtatgccag ccctcagccc ctggccgtgt
     3181 ccacactgcc cggtgtgggg caggtggccc gcccaggacc caccgctgtg ggcaacggcc
     3241 acatggcagg gcccctgctg cctccaccgc cgccagccca gccgtccgcc actctcccca
     3301 gtggtgcccc tgccaccaat gggcccccca caaccgactc ggcccacggg ctgcagatgc
     3361 tgcggaccat tggcgtgggg aagtatgagt tcaccgaccc ggggcacccc agagaaatgt
     3421 tgaaggaatt gaaccagcaa cgcagagcga aagcgtttac agacctgaaa attgttgttg
     3481 aaggcagaga gtttgaagtc caccaaaatg ttctagcttc ctgcagcttg tatttcaagg
     3541 acctgattca aaggtccgtg caagacagcg gccagggcgg ccgggagaag ctggagctcg
     3601 tcctgtcgaa cctgcaggca gacgtcctgg agttgctgct ggagtttgtc tacacgggct
     3661 ccctggtcat cgactcggcc aacgccaaga cactgctgga ggcggccagc aagttccagt
     3721 tccacacctt ctgcaaagtc tgcgtgtcct ttctcgagaa gcagctgacg gccagcaact
     3781 gcctgggcgt gctggccatg gccgaggcca tgcagtgcag cgagctctac cacatggcca
     3841 aggccttcgc gctgcagatc ttccccgagg tggccgccca ggaggagatc ctcagcatct
     3901 ccaaggacga cttcatcgcc tacgtctcca acgacagcct caacaccaag gctgaggagc
     3961 tggtgtacga gacagtcatc aagtggatca agaaggaccc cgcgacacgc acacagtacg
     4021 cggctgagct cctggccgtg gtccgcctcc ccttcatcca ccccagctac ctgctcaatg
     4081 tggttgacaa tgaagagctg atcaagtcat cagaagcctg ccgggacctg gtgaacgagg
     4141 ccaaacgcta ccatatgctg ccccacgccc gccaggagat gcagacgccc cgaacccggc
     4201 cgcgcctctc tgcaggtgtg gctgaggtca tcgtcttggt tgggggccgt cagatggtgg
     4261 ggatgaccca gcgctcgctg gtggccgtca cctgctggaa cccgcagaac aacaagtggt
     4321 accccttggc ctcgctgccc ttctatgacc gcgagttctt cagtgtagtg agtgcagggg
     4381 acaacatcta cctctcaggt gggatggaat caggggtgac gctggctgat gtctggtgct
     4441 acatgtccct gcttgataac tggaacctcg tctccagaat gacagtcccc cgctgtcggc
     4501 acaatagcct cgtctacgat gggaagattt acaccctcgg gggacttggc gtggcaggca
     4561 acgtggacca cgtggagagg tacgacacca tcaccaacca atgggaggcg gtggcccctc
     4621 tgcccaaggc agtacactct gctgcagcca cagtgtgtgg cggcaagatc tacgtgtttg
     4681 gtggggtgaa cgaggcaggc cgagctgccg gcgtcctcca gtcttacgtt cctcagacca
     4741 acacgtggag cttcatcgag tccccaatga ttgacaacaa gtatgccccc gctgtcacgc
     4801 tcaatggctt cgttttcatc ctgggcgggg cttatgccag agctaccacc atctacgacc
     4861 ctgagaaagg aaacattaag gcgggcccaa acatgaacca ctctcgccag ttctgcagtg
     4921 ctgtggtgct tgatggcaag atttatgcaa ctggaggtat tgtcagcagt gaagggcccg
     4981 cgctgggcaa catggaggcc tacgagccca caaccaacac atggaccctc ctcccccaca
     5041 tgccctgccc tgtgttcaga cacggctgcg tcgtgataaa gaaatatatt caaagcggcg
     5101 tcgacgacta caaagaccat gacggtgatt ataaagatca tgacatcgac tacaaggatg
     5161 acgatgacaa gggaagcgga gccactaact tctccctgtt gaaacaagca ggggatgtcg
     5221 aagagaatcc cgggccagaa ttcgccacca tggtgagcaa gggcgaggag ctgttcaccg
     5281 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt
     5341 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca
     5401 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt
     5461 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg
     5521 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg
     5581 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact
     5641 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg
     5701 tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca
     5761 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg
     5821 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag
     5881 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca
     5941 ctctcggcat ggacgagctg tacaagtaag gatccaatca acctctggat tacaaaattt
     6001 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     6061 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     6121 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     6181 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     6241 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     6301 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     6361 tgtcggggaa gctgacgtcc tttccatggc tgctcgcctg tgttgccacc tggattctgc
     6421 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     6481 gcctgctgcc ggctctgcgg cctcttccgc gtctccgcct tcgccctcag acgagtcgga
     6541 tctccctttg gccgcctccc cgcctggtac ctttaagacc aatgacttac aaggcagctg
     6601 tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt cactcccaac
     6661 gaaaataaga tctgcttttt gcttgtactg ggtctctctg gttagaccag atctgagcct
     6721 gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag
     6781 tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac
     6841 ccttttagtc agtgtggaaa atctctagca gtagtagttc atgtcatctt attattcagt
     6901 atttataact tgcaaagaaa tgaatatcag agagtgagag gaacttgttt attgcagctt
     6961 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     7021 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggctctagc
     7081 tatcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     7141 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     7201 aggctttttt ggaggcctag acttttgcag agacggccca aattcgtaat catggtcata
     7261 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     7321 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     7381 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     7441 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     7501 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     7561 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     7621 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     7681 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     7741 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     7801 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     7861 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     7921 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     7981 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     8041 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac
     8101 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     8161 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     8221 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     8281 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     8341 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     8401 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     8461 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     8521 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     8581 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     8641 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     8701 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     8761 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     8821 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     8881 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     8941 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     9001 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     9061 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     9121 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     9181 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     9241 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     9301 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     9361 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     9421 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc
     9481 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     9541 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     9601 cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca
     9661 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgccattc
     9721 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     9781 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
     9841 ccagtcacga cgttgtaaaa cgacggccag tgccaagctg
//
