The CRISPR/Cas9 system is widely used for targeted gene editing. It is engineered from the prokaryotic immune system.
Details
- Bacterial genomes contain a CRISPR locus with the following elements:
- Clustered regularly interspaced short palindromic repeats (where the acronym CRISPR is derived)
- Trans-activating RNA (tracRNA)
- The Cas9 enzyme
- The combination of tracRNA, Cas9, and CRISPR RNA (crRNA) are used to protect against viral genomes with a complementary sequence to the crRNA
- Cas9 scans DNA sequence for protospacer adjacent motif (PAM), three to five nucleotides long. Different Cas9 enzymes recognize different PAMs
- At the PAM site, the helix is unwound and compared to crRNA. Cas9 introduces a double-stranded cut if there is a match
- Autoimmunogenicity is avoided by the bacterium by having zero copies of the PAM in the CRISPR locus
- Engineered versions of this mechanism combine the tracRNA and crRNA into a single guide RNA (sgRNA)
The therapeutic Casgevy which uses this technology was approved in 2023 to treat Sickle-cell anemia.