LOCUS       V045204                 9220 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V045204
VERSION     V045204
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..4439
                     /label="KLHL13(NM_033495)"
                     /note="KLHL13(NM_033495)"
                     /gene="KLHL13"
     CDS             4446..4511
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             4521..4577
                     /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             4590..5309
                     /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    5316..5903
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5977..6210
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6282..6403
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6422..6557
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6595..6611)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6619..6635
                     /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(6643..6673)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6688..6709
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6997..7585)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7756..8616)
                     /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(8617..8721)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9195..9211
                     /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 caccatgcca ttgaaatgga aaacgagctc tcctgctatc tggaaattcc
     2521 cagttcctgt gcttaaaaca tccaggtcaa ctccactttc tccagcatac atatctctcg
     2581 tggaagagga agaccaacac atgaaattgt cccttggagg cagcgaaatg ggcctctcat
     2641 cccatttgca gtcttccaag gcaggaccta cacgcatctt taccagcaat acccacagtt
     2701 ctgtggtgtt acagggcttt gaccagcttc gacttgaagg attgctttgt gatgtgaccc
     2761 tgatgccagg tgacacagat gatgctttcc ctgtgcatag agtcatgatg gcatctgcta
     2821 gtgattactt caaggctatg ttcacaggtg gaatgaaaga acaagattta atgtgcatta
     2881 aacttcatgg tgtgagcaaa gtcggtctaa ggaaaattat tgatttcatt tatactgcaa
     2941 agctttctct taatatggac aaccttcaag acacgctgga agctgccagt ttcctacaga
     3001 ttctgccagt tttggacttc tgtaaagtgt ttctcatatc tggggtcact ttagacaact
     3061 gtgttgaagt tggacggatt gccaacacct acaatctaac cgaagtggat aaatacgtta
     3121 acagtttcgt cttgaagaat tttcctgcat tgctgagcac aggggagttc ttgaaactcc
     3181 cttttgagcg tcttgccttc gtgctttcca gtaatagcct taagcactgt actgaacttg
     3241 agctctttaa ggctacctgt cgttggcttc gcctggaaga gcctcggatg gactttgctg
     3301 caaaattaat gaagaacata cgatttccac tgatgacacc acaggagctc attaattacg
     3361 tgcaaacggt ggatttcatg agaactgaca atacttgtgt gaatttgctt ttggaagcca
     3421 gcaattacca aatgatgcca tatatgcagc cagttatgca gtcagacagg actgccatta
     3481 ggtctgacac cactcacttg gttacactag gaggagtgct gaggcagcag ctggttgtca
     3541 gtaaggaatt gcgcatgtat gatgaaaagg cccatgagtg gaaatcgtta gcccccatgg
     3601 atgccccaag gtaccagcat ggcatcgccg tcattggaaa ttttctctat gtggttggcg
     3661 gacagagtaa ttatgataca aaaggaaaaa cggcagttga tacagtcttc agatttgatc
     3721 ctcgatacaa taaatggatg caagttgcat ctttaaatga aaagcgcacc ttcttccacc
     3781 taagtgccct caaaggatat ctgtatgcag ttggtgggcg aaatgcagca ggtgaactgc
     3841 ccacagtaga atgttacaat ccaagaacaa atgaatggac ctatgttgcc aaaatgagtg
     3901 agccccacta tggccatgct ggaactgtgt atggaggagt gatgtatatt tcaggaggaa
     3961 ttactcatga tactttccaa aaggagctca tgtgctttga ccctgatact gacaaatgga
     4021 tccagaaggc gccaatgacc actgtcagag gtctgcattg catgtgtaca gtgggagaaa
     4081 ggctctatgt cattggtggc aatcacttca gaggaacaag tgattatgat gatgtcctaa
     4141 gctgtgaata ctattcacct atccttgacc agtggacccc aattgctgcc atgttaagag
     4201 ggcagagtga tgttggggtc gctgtcttcg aaaataaaat ctatgtggtt ggggggtatt
     4261 cttggaataa tcgttgtatg gtagagatag tgcagaaata tgatccagat aaagatgaat
     4321 ggcataaggt ttttgatctg ccagaatccc ttggtggcat tcgtgcttgc acactcacag
     4381 tttttccacc agaagaaacc acaccatcac cttctagaga gtcccctctt tctgcacctg
     4441 tcgacgacta caaagaccat gacggtgatt ataaagatca tgacatcgac tacaaggatg
     4501 acgatgacaa gggaagcgga gccactaact tctccctgtt gaaacaagca ggggatgtcg
     4561 aagagaatcc cgggccagaa ttcgccacca tggtgagcaa gggcgaggag ctgttcaccg
     4621 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt
     4681 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca
     4741 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt
     4801 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg
     4861 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg
     4921 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact
     4981 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg
     5041 tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca
     5101 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg
     5161 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag
     5221 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca
     5281 ctctcggcat ggacgagctg tacaagtaag gatccaatca acctctggat tacaaaattt
     5341 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     5401 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     5461 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     5521 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     5581 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     5641 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     5701 tgtcggggaa gctgacgtcc tttccatggc tgctcgcctg tgttgccacc tggattctgc
     5761 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     5821 gcctgctgcc ggctctgcgg cctcttccgc gtctccgcct tcgccctcag acgagtcgga
     5881 tctccctttg gccgcctccc cgcctggtac ctttaagacc aatgacttac aaggcagctg
     5941 tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt cactcccaac
     6001 gaaaataaga tctgcttttt gcttgtactg ggtctctctg gttagaccag atctgagcct
     6061 gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag
     6121 tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac
     6181 ccttttagtc agtgtggaaa atctctagca gtagtagttc atgtcatctt attattcagt
     6241 atttataact tgcaaagaaa tgaatatcag agagtgagag gaacttgttt attgcagctt
     6301 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     6361 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggctctagc
     6421 tatcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     6481 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     6541 aggctttttt ggaggcctag acttttgcag agacggccca aattcgtaat catggtcata
     6601 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     6661 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     6721 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     6781 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     6841 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     6901 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     6961 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     7021 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     7081 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     7141 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     7201 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     7261 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     7321 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     7381 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac
     7441 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     7501 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     7561 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     7621 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     7681 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     7741 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     7801 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     7861 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     7921 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     7981 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     8041 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     8101 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     8161 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     8221 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     8281 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     8341 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     8401 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     8461 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     8521 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     8581 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     8641 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     8701 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     8761 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc
     8821 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     8881 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     8941 cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca
     9001 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgccattc
     9061 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     9121 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
     9181 ccagtcacga cgttgtaaaa cgacggccag tgccaagctg
//
