LOCUS       V044967                 9382 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V044967
VERSION     V044967
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..4601
                     /label="KLHL5(NM_015990)"
                     /note="KLHL5(NM_015990)"
                     /gene="KLHL5"
     CDS             4608..4673
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             4683..4739
                     /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             4752..5471
                     /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    5478..6065
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6139..6372
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6444..6565
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6584..6719
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6757..6773)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6781..6797
                     /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(6805..6835)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6850..6871
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7159..7747)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7918..8778)
                     /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(8779..8883)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9357..9373
                     /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 caccatgtct ggttctcgta aagagtttga tgtgaaacag attttgaaaa
     2521 tcagatggag gtggtttggt catcaagcat catctcctaa ttctacagtt gacagccagc
     2581 agggagaatt ttggaaccga ggacagactg gagcaaacgg tgggagaaag tttttagatc
     2641 catgtagcct acaattgcct ttggcttcaa ttggttaccg aaggtccagc caactggatt
     2701 ttcagaattc accttcttgg ccaatggcat ccacctctga agtccctgca tttgagttta
     2761 cagcagaaga ttgtggcggt gcacattggc tggatagacc agaagtggat gatggcacta
     2821 gtgaagaaga aaatgaatct gattccagtt catgcaggac ttccaatagt agtcagacat
     2881 tatcatcctg tcatactatg gagccatgta catcagatga atttttccaa gcccttaatc
     2941 atgccgagca aacatttaaa aaaatggaaa actatttgag acataaacag ttgtgtgatg
     3001 taattttagt cgctggtgat cgcagaattc cagctcacag attggtgctc tcctctgtct
     3061 cagactattt tgctgccatg tttactaatg atgtcagaga ggcaagacaa gaagaaataa
     3121 aaatggaagg tgtagaacca aattcgttgt ggtccttgat ccagtatgct tatacaggcc
     3181 gccttgaatt aaaagaagat aatattgagt gcctgttatc tacagcttgc cttcttcagc
     3241 tttcacaggt tgtagaagca tgctgtaagt ttttaatgaa acagcttcat ccatccaact
     3301 gtcttggaat tcgttctttt gctgatgccc aaggttgtac agatttgcat aaagtggctc
     3361 acaattatac tatggagcat ttcatggaag taatcagaaa ccaggaattt gtattattac
     3421 cagccagcga aattgcaaag ctcttggcta gtgatgacat gaacattcct aatgaggaga
     3481 caatattgaa tgcacttctt acttgggtcc gtcatgattt ggaacagaga cggaaagatc
     3541 taagtaaact tttggcttat attaggctac ctcttcttgc accacagttc ctggcagaca
     3601 tggaaaataa tgtacttttt cgggatgata tagaatgtca gaaactcatt atggaagcaa
     3661 tgaagtacca tttattacca gagagacgac ccatgttaca aagtcctcgg acaaaaccta
     3721 ggaagtcaac tgttggtaca ttatttgcag ttgggggaat ggattcaaca aaaggagcaa
     3781 caagcattga aaagtatgat ctccgtacaa atatgtggac tccagtagca aatatgaatg
     3841 ggaggaggct acagttcggt gttgcagtgc tagatgacaa actgtatgtg gttggaggaa
     3901 gagatggact gaagactttg aatactgtag agtgctacaa ccccaaaaca aaaacttgga
     3961 gtgtgatgcc acctatgtcc acacatagac atggccttgg tgtggctgta ctggaaggtc
     4021 ccatgtatgc cgtaggagga catgatggct ggagctatct gaacacagtg gaaagatggg
     4081 accctcaggc tcgccagtgg aattttgttg ccactatgtc tacccctagg agtacagtag
     4141 gtgtggcagt actaagtgga aaactttatg cagttggtgg tcgtgatgga agttcttgtc
     4201 tcaaatcagt agaatgtttt gatcctcata ctaataagtg gacactgtgt gcacagatgt
     4261 caaaaaggag aggtggcgta ggagtgacga cctggaatgg actgctgtat gctatagggg
     4321 ggcacgatgc tcccgcatcc aacttgactt ccagactctc agactgtgtg gaaagatatg
     4381 atcccaaaac agacatgtgg actgcagtag catccatgag catcagcaga gatgcagtgg
     4441 gggtctgttt acttggtgat aagttatatg ctgttggggg gtatgatgga caggcatacc
     4501 ttaatactgt ggaggcttat gatccccaga caaatgagtg gacccaggtt gctccactgt
     4561 gcctaggaag agctggagct tgtgttgtga ctgtaaaatt agtcgacgac tacaaagacc
     4621 atgacggtga ttataaagat catgacatcg actacaagga tgacgatgac aagggaagcg
     4681 gagccactaa cttctccctg ttgaaacaag caggggatgt cgaagagaat cccgggccag
     4741 aattcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
     4801 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
     4861 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     4921 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     4981 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     5041 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
     5101 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     5161 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca
     5221 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg
     5281 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     5341 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg
     5401 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     5461 tgtacaagta aggatccaat caacctctgg attacaaaat ttgtgaaaga ttgactggta
     5521 ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc
     5581 atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt
     5641 ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg
     5701 ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt
     5761 tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct
     5821 ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg aagctgacgt
     5881 cctttccatg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct
     5941 acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc
     6001 ggcctcttcc gcgtctccgc cttcgccctc agacgagtcg gatctccctt tggccgcctc
     6061 cccgcctggt acctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt
     6121 taaaagaaaa ggggggactg gaagggctaa ttcactccca acgaaaataa gatctgcttt
     6181 ttgcttgtac tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac
     6241 tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg
     6301 cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga
     6361 aaatctctag cagtagtagt tcatgtcatc ttattattca gtatttataa cttgcaaaga
     6421 aatgaatatc agagagtgag aggaacttgt ttattgcagc ttataatggt tacaaataaa
     6481 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     6541 tgtccaaact catcaatgta tcttatcatg tctggctcta gctatcccgc ccctaactcc
     6601 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     6661 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     6721 agacttttgc agagacggcc caaattcgta atcatggtca tagctgtttc ctgtgtgaaa
     6781 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     6841 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     6901 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     6961 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     7021 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     7081 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     7141 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     7201 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     7261 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     7321 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     7381 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     7441 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     7501 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     7561 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     7621 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     7681 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     7741 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     7801 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     7861 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     7921 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     7981 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     8041 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     8101 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     8161 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     8221 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     8281 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     8341 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     8401 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     8461 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     8521 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     8581 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     8641 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     8701 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     8761 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     8821 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     8881 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
     8941 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg
     9001 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     9061 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
     9121 taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc
     9181 gcacagatgc gtaaggagaa aataccgcat caggcgccat tcgccattca ggctgcgcaa
     9241 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg
     9301 atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa
     9361 aacgacggcc agtgccaagc tg
//
