LOCUS       V044843                 9022 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V044843
VERSION     V044843
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..4241
                     /label="KLHL25(NM_022480)"
                     /note="KLHL25(NM_022480)"
                     /gene="KLHL25"
     CDS             4248..4313
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             4323..4379
                     /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             4392..5111
                     /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    5118..5705
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5779..6012
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6084..6205
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6224..6359
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6397..6413)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6421..6437
                     /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(6445..6475)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6490..6511
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6799..7387)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7558..8418)
                     /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(8419..8523)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     8997..9013
                     /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 caccatgtcg gtcagtgtcc atgagacccg caagtcgcgg agcagcacgg
     2521 ggtccatgaa cgtcaccctc ttccacaagg cctcccaccc ggactgtgtg ctggcccacc
     2581 tcaacacgct tcgcaagcac tgcatgttca ccgacgtcac actctgggcg ggcgaccgtg
     2641 ccttcccctg tcaccgtgcc gtgctggccg cctctagccg ctattttgag gccatgttca
     2701 gccatggcct tcgggagagc cgggatgaca ctgtcaactt ccaggacaac ctgcacccgg
     2761 aggtgctgga gctgctgctg gactttgcct actcctcacg catcgccatc aacgaggaga
     2821 acgctgagtc actgctggag gcaggcgaca tgctgcagtt ccacgatgtg cgggatgctg
     2881 ccgccgagtt cctggagaag aaccttttcc cctccaactg cctgggcatg atgctgctct
     2941 cggacgccca ccagtgccgc cggctgtatg agttctcctg gcgcatgtgc ctggtgcact
     3001 ttgagacggt gaggcagagc gaggacttca acagcctgtc caaggacaca ctgctggacc
     3061 tcatctcgag tgatgagctg gagaccgagg acgagcgggt ggtcttcgag gccatcctcc
     3121 agtgggtgaa gcacgacctg gagccacgga aggtccactt gcccgagctc ctccgcagcg
     3181 tgcgtctggc cttgctgccg tccgactgcc tgcaggaggc cgtctccagc gaggccctcc
     3241 tcatggcaga cgagcgcacc aagcttatca tggatgaggc cctgcgctgc aagaccagga
     3301 tcctgcagaa tgatggcgtg gtcaccagcc cctgtgcccg gccacgcaag gcgggccaca
     3361 cgctactcat cctggggggc cagaccttca tgtgtgacaa gatctaccag gtggaccaca
     3421 aggccaagga gatcatcccc aaggccgacc tgcccagccc ccggaaggag ttcagcgcct
     3481 cagcgatcgg ctgcaaggtc tatgtgacgg ggggcagggg ctccgagaac ggggtctcca
     3541 aggatgtctg ggtgtacgac accgtacatg aggaatggtc caaggcggcg cccatgctga
     3601 ttgcccgctt tggccatggc tcagctgagc tggagaactg cctctatgtg gtggggggac
     3661 acacatccct ggcaggggtc ttcccggcct cgccttctgt ctccctgaaa caagtggaga
     3721 aatacgaccc tggggccaac aagtggatga tggtggcccc cttgcgggat ggcgtcagca
     3781 atgccgcagt ggtgagtgcc aagctgaagc tctttgtttt cggaggaacc agcatccacc
     3841 gggacatggt gtccaaggtc cagtgctatg acccctcgga gaacaggtgg acgatcaagg
     3901 ccgagtgccc ccagccttgg cggtacacag ccgctgccgt cctgggcagc cagatcttca
     3961 tcatgggagg tgacacggaa ttcacagccg cctcggccta ccgctttgac tgtgagacca
     4021 accagtggac gcggattggg gacatgactg ccaagcgcat gtcctgccat gccctggctt
     4081 ccggcaacaa gctctatgtg gtcgggggct actttgggac ccagaggtgt aagactctgg
     4141 actgctatga ccccacttca gatacatgga actgcatcac cacagtgccc tactcactta
     4201 tccccacggc ctttgtcagc acctggaagc acctgcccgc ggtcgacgac tacaaagacc
     4261 atgacggtga ttataaagat catgacatcg actacaagga tgacgatgac aagggaagcg
     4321 gagccactaa cttctccctg ttgaaacaag caggggatgt cgaagagaat cccgggccag
     4381 aattcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
     4441 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
     4501 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     4561 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     4621 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     4681 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
     4741 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     4801 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca
     4861 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg
     4921 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     4981 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg
     5041 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     5101 tgtacaagta aggatccaat caacctctgg attacaaaat ttgtgaaaga ttgactggta
     5161 ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc
     5221 atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt
     5281 ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg
     5341 ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt
     5401 tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct
     5461 ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg aagctgacgt
     5521 cctttccatg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct
     5581 acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc
     5641 ggcctcttcc gcgtctccgc cttcgccctc agacgagtcg gatctccctt tggccgcctc
     5701 cccgcctggt acctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt
     5761 taaaagaaaa ggggggactg gaagggctaa ttcactccca acgaaaataa gatctgcttt
     5821 ttgcttgtac tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac
     5881 tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg
     5941 cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga
     6001 aaatctctag cagtagtagt tcatgtcatc ttattattca gtatttataa cttgcaaaga
     6061 aatgaatatc agagagtgag aggaacttgt ttattgcagc ttataatggt tacaaataaa
     6121 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     6181 tgtccaaact catcaatgta tcttatcatg tctggctcta gctatcccgc ccctaactcc
     6241 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     6301 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     6361 agacttttgc agagacggcc caaattcgta atcatggtca tagctgtttc ctgtgtgaaa
     6421 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     6481 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     6541 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     6601 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     6661 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     6721 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     6781 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     6841 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     6901 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     6961 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     7021 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     7081 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     7141 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     7201 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     7261 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     7321 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     7381 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     7441 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     7501 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     7561 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     7621 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     7681 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     7741 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     7801 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     7861 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     7921 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     7981 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     8041 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     8101 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     8161 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     8221 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     8281 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     8341 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     8401 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     8461 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     8521 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
     8581 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg
     8641 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     8701 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
     8761 taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc
     8821 gcacagatgc gtaaggagaa aataccgcat caggcgccat tcgccattca ggctgcgcaa
     8881 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg
     8941 atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa
     9001 aacgacggcc agtgccaagc tg
//
