dCas9::GSD is a ligand-inducible RNA-guided nuclease generated by fusing 'dCas9' (a catalytically dead form of the Cas9 endonuclease from Streptococcus pyogenes that contains the amino acid substitutions D10A and H840A) with a nuclear localization signal at both the N-terminal and C-terminal ends, the 'GeneSwitch' sequence (a truncated ligand-binding domain from the Homo sapiens progesterone receptor), a 15 residue linker (GGGGS(3)) and the nuclease domain of the FokI restriction endonuclease. DNA cleavage is mediated by the FokI nuclease domain, while binding specificity is mediated by the dCas9 sequence, which requires a guide RNA (gRNA) to target it to a genomic target site of interest. The gRNA is composed of a ~20 nucleotide 'spacer' sequence that defines the genomic target site, plus additional sequence required for dCas9 protein binding (PMID:23287722, PMID:23287718). DNA cleavage by FokI nuclease domains is strictly dimerization dependent, thus DNA cleavage by dCas9::GSD requires the use of two gRNAs that are ~15-25bp apart (PMID:24770324). These requirements effectively extend the length of the target site for dCas9::GSD compared to wild-type Cas9 endonuclease, which is expected to increase the specificity of DNA cleavage (PMID:24770324). In addition, the presence of the GeneSwitch ligand-binding domain together with the 15 residue linker means that FokI domain dimerization and DNA cleavage can only occur in the presence of anti-progestin mifepristone (RU486) (FBrf0245398).