Table 1.

A list of selected Cas9 enzymes and their characteristics.

Cas9 Enzyme	Source Organism	PAM Site (5′-3′)	Size (cDNA, Protein)	Properties	Ref/s
SpCas9	Streptococcus pyogenes	NGG	4.10 kbp, 1,368 aa	Most widely used Cas9; common PAM site, but also more susceptible to off-target effects, multiple variants available	[30,38]
SaCas9	Staphylococcus aureus	NNGRRT	3.16 kbp, 1,053 aa	Small Cas9 with high specificity and efficiency	[39]
BlatCas9	Brevibacillus laterosporus	NNNNCNDD	3.28 kbp, 1,092 aa	Small Cas9 with broad targeting range	[40]
CdCas9	Corynebacterium diphtheriae	NNRHHHY	3.25 kbp, 1,084 aa	Diverse PAM sequence recognition, more robust DNA cleavage activity compared to traditional Cas9 orthologues	[41]
CjCas9	Campylobacter jejuni	NNNNACA	2.95 kbp, 984 aa	Smallest available Cas9, lack of a protospacer flanking sequence allows for greater target flexibility, shown to target RNA as well	[42,43]
FnCas9	Francisella novicida	NGG	4.89 kbp, 1,629 aa	High binding specificity and low off-target effect compared to SpCas9 while having similar PAM site, size may deter delivery	[44]
GeoCas9	Geobacillus stearothermophilus	CRAA	3.26 kbp, 1,087 aa	Small thermotolerant Cas9 active in temperatures up to 70 °C, shown to have improved ribonucleoprotein delivery	[45]
NmCas9	Neisseria meningitidis	NNNNGATT	3.25 kbp, 1,082 aa	Small Cas9 with high specificity, lower off-target effect compared to SpCas9	[46,47]
ScCas9	Streptococcus canis	NNG	4.161 kbp, 1,386 aa	Homologous protein to SpCas9 with higher levels of specificity, size may deter delivery	[48]
St1Cas9	Streptococcus thermophilus	NNAGAAW	3.36 kbp, 1,121 aa	Small Cas9 that can induce error-prone NHEJ-mediated targeted mutagenesis as efficiently as SpCas9	[49]

Abbreviations: PAM, protospacer adjacent motif; NHEJ, non-homologous end joining.