LOCUS       sequence-177915-        4404 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expression of dead/inactive increased fidelity eSpCas9 
            (1.1)-VP64-6xHis in bacterial cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 4404)
  AUTHORS   Kulcsar PI, Talas A, Huszar K, Ligeti Z, Toth E, Weinhardt N, Fodor 
            E, Welker E
  TITLE     Crossing enhanced and high fidelity SpCas9 nucleases to optimize 
            specificity and cleavage.
  JOURNAL   Genome Biol. 2017 Oct 6;18(1):190. doi: 10.1186/s13059-017-1318-8.
  PUBMED    28985763
REFERENCE   2  (bases 1 to 4404)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Genome 
            Biol."; date: "2017-10-6"; volume: "18(1):190. doi"; pages: " 
            10.1186/s13059-017-1318-8"
COMMENT     Sequence Label: pET-deSpCas9-VP64-6xHis
FEATURES             Location/Qualifiers
     source          1..4404
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             4111..4131
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4138..4203
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             4234..4383
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             4384..4401
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 atggataaga aatactcaat aggcttagct atcggcacaa atagcgtcgg atgggcggtg
       61 atcactgatg aatataaggt tccgtctaaa aagttcaagg ttctgggaaa tacagaccgc
      121 cacagtatca aaaaaaatct tataggggct cttttatttg acagtggaga gacagcggaa
      181 gcgactcgtc tcaaacggac agctcgtaga aggtatacac gtcggaagaa tcgtatttgt
      241 tatctacagg agattttttc aaatgagatg gcgaaagtag atgatagttt ctttcatcga
      301 cttgaagagt cttttttggt ggaagaagac aagaagcatg aacgtcatcc tatttttgga
      361 aatatagtag atgaagttgc ttatcatgag aaatatccaa ctatctatca tctgcgaaaa
      421 aaattggtag attctactga taaagcggat ttgcgcttaa tctatttggc cttagcgcat
      481 atgattaagt ttcgtggtca ttttttgatt gagggagatt taaatcctga taatagtgat
      541 gtggacaaac tatttatcca gttggtacaa acctacaatc aattatttga agaaaaccct
      601 attaacgcaa gtggagtaga tgctaaagcg attctttctg cacgattgag taaatcaaga
      661 cgattagaaa atctcattgc tcagctcccc ggtgagaaga aaaatggctt atttgggaat
      721 ctcattgctt tgtcattggg tttgacccct aattttaaat caaattttga tttggcagaa
      781 gatgctaaat tacagctttc aaaagatact tacgatgatg atttagataa tttattggcg
      841 caaattggag atcaatatgc tgatttgttt ttggcagcta agaatttatc agatgctatt
      901 ttactttcag atatcctaag agtaaatact gaaataacta aggctcccct atcagcttca
      961 atgattaaac gctacgatga acatcatcaa gacttgactc ttttaaaagc tttagttcga
     1021 caacaacttc cagaaaagta taaagaaatc ttttttgatc aatcaaaaaa cggatatgca
     1081 ggttatattg atgggggagc tagccaggaa gagttctaca agttcatcaa gcccatcctg
     1141 gaaaagatgg acggcaccga ggaactgctc gtgaagctga acagagagga cctgctgcgg
     1201 aagcagcgga ccttcgacaa cggcagcatc ccccaccaga tccacctggg agagctgcac
     1261 gccattctgc ggcggcagga agatttttac ccattcctga aggacaaccg ggaaaagatc
     1321 gagaagatcc tgaccttccg catcccctac tacgtgggcc ctctggccag gggaaacagc
     1381 agattcgcct ggatgaccag aaagagcgag gaaaccatca ccccctggaa cttcgaggaa
     1441 gtggtggaca agggcgcttc cgcccagagc ttcatcgagc ggatgaccaa cttcgataag
     1501 aacctgccca acgagaaggt gctgcccaag cacagcctgc tgtacgagta cttcaccgtg
     1561 tataacgagc tgaccaaagt gaaatacgtg accgagggaa tgagaaagcc cgccttcctg
     1621 agcggcgagc agaaaaaggc catcgtggac ctgctgttca agaccaaccg gaaagtgacc
     1681 gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt gcttcgactc cgtggaaatc
     1741 tccggcgtgg aagatcggtt caacgcctcc ctgggcacat accacgatct gctgaaaatt
     1801 atcaaggaca aggacttcct ggacaatgag gaaaacgagg acattctgga agatatcgtg
     1861 ctgaccctga cactgtttga ggacagagag atgatcgagg aacggctgaa aacctatgcc
     1921 cacctgttcg acgacaaagt gatgaagcag ctgaagcggc ggagatacac cggctggggc
     1981 aggctgagcc ggaagctgat caacggcatc cgggacaagc agtccggcaa gacaatcctg
     2041 gatttcctga agtccgacgg cttcgccaac agaaacttca tgcagctgat ccacgacgac
     2101 agcctgacct ttaaagagga catccagaaa gcccaggtgt ccggccaggg cgatagcctg
     2161 cacgagcaca ttgccaatct ggccggcagc cccgccatta agaagggcat cctgcagaca
     2221 gtgaaggtgg tggacgagct cgtgaaagtg atgggccggc acaagcccga gaacatcgtg
     2281 atcgaaatgg ccagagagaa ccagaccacc cagaagggac agaagaacag ccgcgagaga
     2341 atgaagcgga tcgaagaggg catcaaagag ctgggcagcc agatcctgaa agaacacccc
     2401 gtggaaaaca cccagctgca gaacgagaag ctgtacctgt actacctgca gaatgggcgg
     2461 gatatgtacg tggaccagga actggacatc aaccggctgt ccgactacga tgtggacgcc
     2521 atcgtgcctc agagctttct ggccgacgac tccatcgaca acaaggtgct gaccagaagc
     2581 gacaagaacc ggggcaagag cgacaacgtg ccctccgaag aggtcgtgaa gaagatgaag
     2641 aactactggc ggcagctgct gaacgccaag ctgattaccc agagaaagtt cgacaatctg
     2701 accaaggccg agagaggcgg cctgagcgaa ctggataagg ccggcttcat caagagacag
     2761 ctggtggaaa cccggcagat cacaaagcac gtggcacaga tcctggactc ccggatgaac
     2821 actaagtacg acgagaatga caagctgatc cgggaagtga aagtgatcac cctgaagtcc
     2881 aagctggtgt ccgatttccg gaaggatttc cagttttaca aagtgcgcga gatcaacaac
     2941 taccaccacg cccacgacgc ctacctgaac gccgtcgtgg gaaccgccct gatcaaaaag
     3001 taccctgcgc tggaaagcga gttcgtgtac ggcgactaca aggtgtacga cgtgcggaag
     3061 atgatcgcca agagcgagca ggaaatcggc aaggctaccg ccaagtactt cttctacagc
     3121 aacatcatga actttttcaa gaccgagatt accctggcca acggcgagat ccggaaggcg
     3181 cctctgatcg agacaaacgg cgaaaccggg gagatcgtgt gggataaggg ccgggatttt
     3241 gccaccgtgc ggaaagtgct gagcatgccc caagtgaata tcgtgaaaaa gaccgaggtg
     3301 cagacaggcg gcttcagcaa agagtctatc ctgcccaaga ggaacagcga taagctgatc
     3361 gccagaaaga aggactggga tccaaaaaaa tatggtggtt ttgatagtcc aacggtagct
     3421 tattcagtcc tagtggttgc taaggtggaa aaagggaaat cgaagaagtt aaaatccgtt
     3481 aaagagttac tagggatcac aattatggaa agaagttcct ttgaaaaaaa tccgattgac
     3541 tttttagaag ctaaaggata taaggaagtt aaaaaagact taatcattaa actacctaaa
     3601 tatagtcttt ttgagttaga aaacggtcgt aaacggatgc tggctagtgc cggagaatta
     3661 caaaaaggaa atgagctggc tctgccaagc aaatatgtga attttttata tttagctagt
     3721 cattatgaaa agttgaaggg tagtccagaa gataacgaac aaaaacaatt gtttgtggag
     3781 cagcataagc attatttaga tgagattatt gagcaaatca gtgaattttc taagcgtgtt
     3841 attttagcag atgccaattt agataaagtt cttagtgcat ataacaaaca tagagacaaa
     3901 ccaatacgtg aacaagcaga aaatattatt catttattta cgttgacgaa tcttggagct
     3961 cccgctgctt ttaaatattt tgatacaaca attgatcgta aacgatatac gtctacaaaa
     4021 gaagttttag atgccactct tatccatcaa tccatcactg gtctttatga aacacgcatt
     4081 gatttgagtc agctaggagg tgacggttct cccaagaaga agaggaaagt ctcgagcgac
     4141 tacaaagacc atgacggtga ttataaagat catgacatcg attacaagga tgacgatgac
     4201 aaggctgcag gaggcggtgg aagcgggcgc gccgacgcgc tggacgattt cgatctcgac
     4261 atgctgggtt ctgatgccct cgatgacttt gacctggata tgttgggaag cgacgcattg
     4321 gatgactttg atctggacat gctcggctcc gatgctctgg acgatttcga tctcgatatg
     4381 ttacatcacc accaccatca ctaa
//
