LOCUS       V044886                 9082 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V044886
VERSION     V044886
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..4301
                     /label="KLHL20(NM_014458)"
                     /note="KLHL20(NM_014458)"
                     /gene="KLHL20"
     CDS             4308..4373
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             4383..4439
                     /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             4452..5171
                     /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    5178..5765
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5839..6072
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6144..6265
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6284..6419
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6457..6473)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6481..6497
                     /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(6505..6535)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6550..6571
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6859..7447)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7618..8478)
                     /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(8479..8583)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9057..9073
                     /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 caccatggaa ggaaagccaa tgcgcaggtg taccaacatt cgaccaggag
     2521 agactggaat ggatgtaaca agccgctgca cccttggaga ccccaacaaa ctgccagaag
     2581 gggttcccca acctgcccgc atgccctata tctcagacaa gcaccctcga caaaccttgg
     2641 aagtgattaa ccttctgaga aagcaccggg agctatgtga tgtggtgcta gttgtgggcg
     2701 ccaagaagat atatgcccat cgagtcattt tgtcagcctg tagtccctac ttccgagcta
     2761 tgtttacagg agaattggca gagagccgtc agacagaagt agtgatccga gacattgacg
     2821 agagggctat ggaattactg attgactttg cgtatacctc ccagataaca gtagaagagg
     2881 gcaatgttca gactcttctg ccagctgctt gcctcctcca gctggcagaa atacaggaag
     2941 cctgctgtga attcttaaag agacaattag atccttctaa ctgcctgggc attcgggctt
     3001 ttgctgacac acattcatgt cgtgagttgc taaggatagc agacaagttc acccaacata
     3061 actttcaaga ggtaatggag agtgaagagt tcatgttgct tccagccaat caactcattg
     3121 atataatatc cagtgatgag ctaaacgttc gcagtgaaga acaagtgttc aatgcagtga
     3181 tggcctgggt caaatacagt attcaggaaa gacgtcctca attaccccag gtgctgcagc
     3241 atgttcgttt gcctttgctt agtcccaagt tcctggtcgg cacagtaggc tctgatcccc
     3301 tcatcaaaag tgatgaagaa tgcagagact tggtagatga ggctaaaaac tacctcctat
     3361 tgccgcaaga acgaccacta atgcaaggac caaggacgag accacggaaa cctatccgat
     3421 gtggggaagt actctttgca gttggtggtt ggtgcagtgg agatgccatt tccagtgttg
     3481 aacgatatga tccacagacc aatgaatgga gaatggtggc ttcaatgagc aaaaggagat
     3541 gcggagttgg ggtcagtgtt cttgatgatc tgttatatgc agtaggaggc catgatggat
     3601 cctcttatct caatagtgtt gaaaggtatg accccaaaac aaaccagtgg agcagtgatg
     3661 tggcccctac aagcacctgc aggacaagtg ttggtgtagc agtacttgga ggctttcttt
     3721 atgctgtggg tggccaggat ggtgtgtctt gcctcaacat tgttgagagg tatgatccga
     3781 aggagaacaa gtggactcgg gtagcttcta tgagtaccag aagactaggt gtggctgtgg
     3841 ctgtgttagg agggttctta tatgctgtag gtggctctga cgggacatct cctctcaaca
     3901 cagtggaacg ttacaatcct caggaaaaca gatggcacac tatagcccct atggggaccc
     3961 ggaggaaaca cctaggctgt gcagtatatc aggacatgat ctatgctgta ggaggtagag
     4021 atgacactac agagctgagc agtgctgaga gatacaaccc cagaaccaac cagtggtctc
     4081 cagtggtggc catgacatca cgccgtagtg gagttggcct ggcagtggtc aatggacagc
     4141 tcatggcagt aggaggtttt gatggcacaa catacttgaa gaccatagaa gtttttgatc
     4201 ctgatgccaa tacatggagg ttatatggcg ggatgaatta ccgtcggcta gggggtggcg
     4261 taggagttat taaaatgaca cattgtgaat cccatatttg ggtcgacgac tacaaagacc
     4321 atgacggtga ttataaagat catgacatcg actacaagga tgacgatgac aagggaagcg
     4381 gagccactaa cttctccctg ttgaaacaag caggggatgt cgaagagaat cccgggccag
     4441 aattcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
     4501 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
     4561 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     4621 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     4681 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     4741 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
     4801 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     4861 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca
     4921 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg
     4981 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     5041 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg
     5101 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     5161 tgtacaagta aggatccaat caacctctgg attacaaaat ttgtgaaaga ttgactggta
     5221 ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc
     5281 atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt
     5341 ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg
     5401 ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt
     5461 tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct
     5521 ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg aagctgacgt
     5581 cctttccatg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct
     5641 acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc
     5701 ggcctcttcc gcgtctccgc cttcgccctc agacgagtcg gatctccctt tggccgcctc
     5761 cccgcctggt acctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt
     5821 taaaagaaaa ggggggactg gaagggctaa ttcactccca acgaaaataa gatctgcttt
     5881 ttgcttgtac tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac
     5941 tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg
     6001 cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga
     6061 aaatctctag cagtagtagt tcatgtcatc ttattattca gtatttataa cttgcaaaga
     6121 aatgaatatc agagagtgag aggaacttgt ttattgcagc ttataatggt tacaaataaa
     6181 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     6241 tgtccaaact catcaatgta tcttatcatg tctggctcta gctatcccgc ccctaactcc
     6301 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     6361 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     6421 agacttttgc agagacggcc caaattcgta atcatggtca tagctgtttc ctgtgtgaaa
     6481 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     6541 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     6601 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     6661 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     6721 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     6781 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     6841 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     6901 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     6961 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     7021 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     7081 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     7141 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     7201 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     7261 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     7321 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     7381 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     7441 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     7501 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     7561 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     7621 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     7681 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     7741 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     7801 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     7861 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     7921 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     7981 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     8041 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     8101 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     8161 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     8221 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     8281 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     8341 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     8401 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     8461 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     8521 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     8581 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
     8641 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg
     8701 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     8761 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
     8821 taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc
     8881 gcacagatgc gtaaggagaa aataccgcat caggcgccat tcgccattca ggctgcgcaa
     8941 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg
     9001 atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa
     9061 aacgacggcc agtgccaagc tg
//
