LOCUS       V044859                 9139 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V044859
VERSION     V044859
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..4358
                     /label="KLHL14(NM_020805)"
                     /note="KLHL14(NM_020805)"
                     /gene="KLHL14"
     CDS             4365..4430
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             4440..4496
                     /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             4509..5228
                     /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    5235..5822
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5896..6129
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6201..6322
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6341..6476
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6514..6530)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6538..6554
                     /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(6562..6592)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6607..6628
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6916..7504)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7675..8535)
                     /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(8536..8640)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9114..9130
                     /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 agatccgggg acaggacctc caccttcgac cccagccaca
     2521 gcgacaacct gctgcacggc ctcaacctgc tgtggaggaa gcagctgttt tgcgacgtga
     2581 ccctgacggc ccagggccag cagttccatt gccacaaggc cgtgctggcc tcctgctcgc
     2641 agtacttccg atcgctcttc tccagccacc cccctctcgg gggaggggtc ggcggccagg
     2701 acggcctggg ggcccccaag gaccagcagc agccgccgca gcagcagccg tcacagcagc
     2761 agcagccgcc gccgcaggag gagcccggga ctccttcttc ctcccccgac gacaagctgc
     2821 tgaccagccc ccgggccatc aacaacctgg tgctgcaggg ctgctcgtcc atcgggctgc
     2881 gcctggtgct cgagtacctc tacacggcca acgtgaccct gtccctggac acggtggagg
     2941 aggtgctgtc ggtcagcaag atcctgcaca tcccccaggt caccaagctc tgcgtgcagt
     3001 tcctcaacga ccagatctcg gtgcagaact acaagcaggt gtgcaagatc gccgcgctgc
     3061 acggcctgga ggagaccaag aagctggcca acaagtacct ggtggaggat gtgctgctgc
     3121 tcaacttcga ggagatgcgc gccctgctgg actcgctgcc gccccccgtg gagtcggagc
     3181 tggcgctctt ccagatgtcc gtgctgtggc tggagcacga ccgcgagacc cgcatgcagt
     3241 atgcgcctga cctcatgaag cgcctccgct tcgccctcat cccggccccg gagctggtgg
     3301 agcgggtcca gtcagtggat ttcatgcgaa ccgacccggt ctgccagaag ctgctgctgg
     3361 acgccatgaa ctaccacctg atgcccttca ggcagcactg caggcagagc ctggccagca
     3421 gaattcgctc taacaagaaa atgctgttat tggttggagg gctgcctcct ggaccggacc
     3481 ggctccccag caatttggtt cagtattacg acgatgaaaa gaagacatgg aaaatactca
     3541 caattatgcc atacaacagt gcccaccact gcgttgtgga ggtggaaaac ttcttgttcg
     3601 tgttgggtgg agaggaccag tggaatccga atggaaaaca cagtacaaat tttgtcagcc
     3661 gatatgatcc tcgatttaat agctggattc aacttccacc catgcaggaa agaagagcca
     3721 gtttctatgc atgtcggttg gacaagcatt tatacgtaat tggtggaagg aatgaaactg
     3781 gctacttgtc cagcgtggag tgctataacc tagaaacgaa tgaatggcgc tatgtgtcct
     3841 ctttgccaca gcctctggcg gctcatgcgg gagcagtgca caatgggaaa atatacattt
     3901 cagggggtgt acacaatgga gaatatgtcc catggctata ttgctatgac ccagtaatgg
     3961 atgtctgggc tcgaaaacaa gatatgaaca caaaacgtgc aattcacact ttggctgtaa
     4021 tgaatgatcg cttgtatgca attggaggaa atcatttgaa aggtttctcc caccttgatg
     4081 taatgcttgt ggaatgctat gacccaaaag gtgaccagtg gaatatactc caaactccca
     4141 ttttggaggg tcgaagtggc cctggctgtg cagtgcttga tgacagcatt taccttgtgg
     4201 gaggctacag ctggagtatg ggggcctaca agtcatctac aatatgctat tgtccagaga
     4261 aaggaacctg gacagaactc gaaggagatg tagcagaacc gttggcaggc cctgcctgtg
     4321 tgacagttat tctgccctct tgtgtaccat acaacaaagt cgacgactac aaagaccatg
     4381 acggtgatta taaagatcat gacatcgact acaaggatga cgatgacaag ggaagcggag
     4441 ccactaactt ctccctgttg aaacaagcag gggatgtcga agagaatccc gggccagaat
     4501 tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     4561 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     4621 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     4681 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     4741 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     4801 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     4861 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     4921 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     4981 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     5041 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     5101 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     5161 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     5221 acaagtaagg atccaatcaa cctctggatt acaaaatttg tgaaagattg actggtattc
     5281 ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg
     5341 ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc
     5401 tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg
     5461 acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg
     5521 ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga
     5581 caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaag ctgacgtcct
     5641 ttccatggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg
     5701 tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc
     5761 ctcttccgcg tctccgcctt cgccctcaga cgagtcggat ctccctttgg ccgcctcccc
     5821 gcctggtacc tttaagacca atgacttaca aggcagctgt agatcttagc cactttttaa
     5881 aagaaaaggg gggactggaa gggctaattc actcccaacg aaaataagat ctgctttttg
     5941 cttgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
     6001 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
     6061 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
     6121 tctctagcag tagtagttca tgtcatctta ttattcagta tttataactt gcaaagaaat
     6181 gaatatcaga gagtgagagg aacttgttta ttgcagctta taatggttac aaataaagca
     6241 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     6301 ccaaactcat caatgtatct tatcatgtct ggctctagct atcccgcccc taactccgcc
     6361 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     6421 ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctaga
     6481 cttttgcaga gacggcccaa attcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     6541 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     6601 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     6661 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     6721 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     6781 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     6841 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     6901 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     6961 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     7021 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     7081 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     7141 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     7201 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     7261 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     7321 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
     7381 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     7441 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     7501 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     7561 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     7621 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     7681 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     7741 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     7801 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     7861 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     7921 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     7981 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     8041 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     8101 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     8161 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     8221 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     8281 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     8341 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     8401 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     8461 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     8521 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     8581 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     8641 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
     8701 ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga
     8761 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     8821 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa
     8881 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca
     8941 cagatgcgta aggagaaaat accgcatcag gcgccattcg ccattcaggc tgcgcaactg
     9001 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
     9061 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
     9121 gacggccagt gccaagctg
//
