Single guide RNAs are used in the CRISPR-Cas9 system to allow the specific targeting of the Cas9 endonuclease. This enables Cas9 to be targeted to a specific location or sequence in the DNA, providing the specificity that is provided with CRISPR gene editing.
An sgRNA consists of the tracer RNA and guide RNA fused together to form the sgRNA. The guide RNA (gRNA) binds to complementary sequences, identifying the target locus / loci of the genome. The tracer RNA (tracrRNA) provides the binding site for Cas9, allowing it to identify the specific location it is to cleave the DNA at. The Cas9 endonuclease recognises the PAM site, at the 3’ end of the guide RNA. It is thought this destabilises Cas9, on recognition of the NGG sequence, inducing it to cleave the DNA. Modified Cas9 nucleases have been developed that are able to identify alternative PAM sites.
Prime editing is a more advanced Cas9-based gene editing technique, enabling the nicking of DNA to reduce the likelihood of non-homologous end joining or other DNA repair processes from occurring. This improves precision, and is a recent advance in gene editing technology.