Genomic DNA sequence complementary to Cas9-associated sgRNA (Protospacer) (Protospacer)

The genomic DNA sequence complementary to a Cas9-associated single guide RNA (sgRNA) is the fundamental target site for CRISPR-Cas9 gene-editing systems. This specific sequence, known as the protospacer, is identified by the Cas9 protein through RNA-DNA hybridization with the sgRNA spacer sequence, a process that requires the presence of a neighboring Protospacer Adjacent Motif (PAM) (Jinek et al., 2012). Once the Cas9-sgRNA complex binds to this target, the Cas9 enzyme's HNH and RuvC nuclease domains induce a site-specific double-strand break (DSB) in the DNA (Hsu et al., 2013). This break triggers cellular DNA repair mechanisms, such as non-homologous end joining (NHEJ), which can be harnessed to knock out genes or regulatory elements, or homology-directed repair (HDR) for precise sequence correction. In therapeutic contexts, such as the treatment of sickle cell disease with exagamglogene autotemcel, the target is often a specific erythroid-specific enhancer (e.g., within the BCL11A gene) to induce fetal hemoglobin production (Frangoul et al., 2021). The precision of targeting this DNA sequence is paramount, as off-target activity at similar genomic sites can lead to unintended mutations or chromosomal rearrangements (Fu et al., 2013). Consequently, this target represents the nexus of modern precision medicine, enabling direct modification of the human genome to treat previously incurable genetic disorders (Gillmore et al., 2021).

Other names

CRISPR target sitesgRNA-complementary DNA sequenceTarget DNA locusRNA-guided DNA targetCas9 target sequence

RNA-guided site-specific DNA cleavage or modification