An overview of currently utilized genome editor technologies based on CRISPR-associated nucleases and their derivatives. DSB-based genome editing: Cas9 and Cas12a nucleases enable efficient gene knockouts and provide limited capability to generate HDR-mediated knockin edits. Base editing: this approach employs a fusion of a Cas9 nickase (nCas9) with nucleobase modifying enzymes. Base editors enable the direct conversion of a single-nucleotide base into another without the need for double-strand breaks. This approach is particularly effective for introducing specific point mutations (A-to-G or C-to-T, and also A-to-C or C-to-G), enabling precise gene correction or the introduction of stop codons for precise gene knockouts. Prime editing: this technology combines a Cas9 nickase with a reverse transcriptase (RT) and uses a prime editing guide RNA (pegRNA) consisting of a Cas9 sgRNA fused to an RT template (RTT) and a primer-binding site (PBS). Nicking the non-target DNA strand enables its extension by RT upon hybridization to the PBS in the pegRNA, thereby copying the RTT sequence into the target locus. Prime editing enables the insertion, deletion, and replacement of short DNA sequences up to several tens of nucleotides. Transcriptional modulators: these tools enable RNA-guided control of gene transcription by targeting a deactivated Cas9 (dCas9) fused with transcriptional modulation domains (such as VP64 or KRAB) to gene promoters. RNA editors: diverging from genome editing, RNA editors utilize RNA-targeting Cas13 nucleases, either for targeted transcript degradation (when catalytically active) or for transcript editing (when rendered catalytically inactive and fused to adenosine deaminases).
