Genomic double-stranded DNA target site (CRISPR-Cas9) (dsDNA target)

Genomic double-stranded DNA (dsDNA) at specific 20-nucleotide protospacer sequences serves as the fundamental target for CRISPR-Cas9-based genome editing therapies. These target sites are defined by their complementarity to a synthetic guide RNA (gRNA) and their proximity to a Protospacer Adjacent Motif (PAM), which is essential for Cas9 protein recognition and binding (Jinek et al., 2012, Science). Once the Cas9-gRNA complex binds to the target dsDNA, the nuclease domains induce a site-specific double-strand break (DSB). This DSB is subsequently repaired by the cell's endogenous machinery, typically via non-homologous end joining (NHEJ) or homology-directed repair (HDR), leading to gene knockout or precise sequence correction (Doudna & Charpentier, 2014, Science). This molecular target is the basis for recently approved therapies like Exagamglogene autotemcel, which targets the BCL11A enhancer to treat sickle cell disease (Frangoul et al., 2021, NEJM). Therapeutic applications focus on correcting pathogenic mutations or modulating gene expression in various tissues, including hematopoietic stem cells and the liver (Gillmore et al., 2021, NEJM). Beyond simple cleavage, modified Cas proteins can target these sequences to perform base editing or prime editing without inducing double-strand breaks (Anzalone et al., 2019, Nature). The specificity of this target is determined by the 20-nt sequence, allowing for highly programmable therapeutic interventions across the human genome.

Other names

ProtospacerCRISPR target sequenceGenomic DNA targetCas9 target siteTarget DNA sequence

RNA-guided, site-specific DNA cleavage followed by endogenous DNA repair mechanisms (NHEJ or HDR) to achieve gene disruption, correction, or insertion.