LOCUS       sequence-107609-         986 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses Fok1 nuclease domain fused to catalytically inactive Cas9 
            DNA-binding domain in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 986)
  AUTHORS   Guilinger JP, Thompson DB, Liu DR
  TITLE     Fusion of catalytically inactive Cas9 to FokI nuclease improves the 
            specificity of genome modification.
  JOURNAL   Nat Biotechnol. 2014 Apr 25. doi: 10.1038/nbt.2909.
  PUBMED    24770324
REFERENCE   2  (bases 1 to 986)
  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: "Nat 
            Biotechnol. 2014 Apr 25. doi: 10.1038/nbt.2909."
COMMENT     Sequence Label: FokI-dCas9
FEATURES             Location/Qualifiers
     source          1..986
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     58..77
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        58..76
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      84..93
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             93..158
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             165..185
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             222..809
                     /codon_start=1
                     /product="nonspecific DNA cleavage domain of the FokI 
                     endonuclease (Li et al., 1992)"
                     /label=FokI cleavage domain
                     /note="must dimerize to be catalytically active"
                     /translation="QLVKSELEEKKSELRHKLKYVPHEYIELIEIARNSTQDRILEMKV
                     MEFFMKVYGYRGKHLGGSRKPDGAIYTVGSPIDYGVIVDTKAYSGGYNLPIGQADEMQR
                     YVEENQTRNKHINPNEWWKVYPSSVTEFKFLFVSGHFKGNYKAQLTRLNHITNCNGAVL
                     SVEELLIGGEMIKAGTLTLEEVRRKFNNGEINF"
ORIGIN
        1 ctatataagc agagctggtt tagtgaaccg tcagatccgc tagagatccg cggccgctaa
       61 tacgactcac tatagggaga gccgccacca tggactacaa agaccatgac ggtgattata
      121 aagatcatga catcgattac aaggatgacg atgacaagat ggcccccaag aagaagagga
      181 aggtgggcat tcaccgcggg gtacctggag gttctggatc ccaactagtc aaaagtgaac
      241 tggaggagaa gaaatctgaa cttcgtcata aattgaaata tgtgcctcat gaatatattg
      301 aattaattga aattgccaga aattccactc aggatagaat tcttgaaatg aaggtaatgg
      361 aattttttat gaaagtttat ggatatagag gtaaacattt gggtggatca aggaaaccgg
      421 acggagcaat ttatactgtc ggatctccta ttgattacgg tgtgatcgtg gatactaaag
      481 cttatagcgg aggttataat ctgccaattg gccaagcaga tgaaatgcaa cgatatgtcg
      541 aagaaaatca aacacgaaac aaacatatca accctaatga atggtggaaa gtctatccat
      601 cttctgtaac ggaatttaag tttttatttg tgagtggtca ctttaaagga aactacaaag
      661 ctcagcttac acgattaaat catatcacta attgtaatgg agctgttctt agtgtagaag
      721 agcttttaat tggtggagaa atgattaaag ccggcacatt aaccttagag gaagtcagac
      781 ggaaatttaa taacggcgag ataaacttta gcggcagcga gactcccggg acctcagagt
      841 ccgccacacc cgaaagtgat aaaaagtatt ctattggttt agctatcggc actaattccg
      901 ttggatgggc tgtcataacc gatgaataca aagtaccttc aaagaaattt aaggtgttgg
      961 ggaacacaga ccgtcattcg attaaa
//
