. 2024 Dec 18;12:156. doi: 10.1186/s40364-024-00701-x

Table 1b.

Characteristics of the different CRISPR systems

Type of CRISPR/Cas system	Subtypes	Organisms	Mechanism of action	Applications
Type II	II-A II-B II-C	• Streptococcus pyogenes • Streptococcus thermophilus • Staphylococcus aureus • Neisseria meningitidis • Campylobacter jejuni	Cas9 creates double-stranded DNA breaks, near the PAM site, which can then be repaired by either non-homologous end joining or homologous recombination with a donor template DNA to create site-specific edits. Type II-A Cas9s generally have high genome editing efficiency, but off-target cleavage at unintended genome sites can be a disadvantage.	− Cancer immunotherapy − Genome-wide screening for identification of cancer genes − Gene therapy − Viral diseases diagnosis and treatment − Tissue regeneration
Type V	V-A (Cpf1) V-B (C2c1)	• Francisella novicida, • Acidaminococcus sp. • Lachnospiraceae sp. • Prevotella sp.	Both subtypes generate staggered DNA breaks distal to the PAM site and enable versatile multiple genomic loci targeting by processing its own guide RNAs.	Cas12 has been engineered as a platform for epigenome editing, and it was recently discovered that Cas12 can indiscriminately chop up single-stranded DNA once activated by a target DNA molecule matching its spacer sequence. This property makes Cas12 a powerful tool for: − Multiplex genome editing − Epigenome editing − Diagnostic of viral diseases The subtype V-B has a significantly smaller size (1108 aa) than subtype V-A (1307 aa), making it a more suitable tool for gene therapy applications that use a viral delivery system. Furthermore, it is very sensitive to mismatches between the guide RNA and target DNA, thus causing less off-target effects.
Type VI	VI-A VI-B VI-C VI-D	• Leptotrichia buccalis • Leptotrichia shahii • Ruminococcus flavefaciens • Bergeyella zoohelcum • Prevotella buccae • Listeria seeligeri	Cas13 targets RNA, not DNA. Once it is activated by a ssRNA sequence complementary to its crRNA spacer, it destroys all nearby RNAs, regardless of their sequence.	Cas13 is an outlier in the CRISPR world because it targets RNA, not DNA. Its applications include: − Specific RNA knock-down or editing without altering genome sequence − Diagnostic of viral diseases (RNA viruses) Subtype VI-B had consistently increased levels of knock-down relative to subtype VI-A. (average of 92.3% versus 40.1%)

Type of CRISPR/Cas system

Subtypes

Organisms

Mechanism of action

Applications

Type II

II-A

II-B

II-C

• Streptococcus pyogenes

• Streptococcus thermophilus

• Staphylococcus aureus

• Neisseria meningitidis

• Campylobacter jejuni

Cas9 creates double-stranded DNA breaks, near the PAM site, which can then be repaired by either non-homologous end joining or homologous recombination with a donor template DNA to create site-specific edits.

Type II-A Cas9s generally have high genome editing efficiency, but off-target cleavage at unintended genome sites can be a disadvantage.

− Cancer immunotherapy

− Genome-wide screening for identification of cancer genes

− Gene therapy

− Viral diseases diagnosis and treatment

− Tissue regeneration

Type V

V-A (Cpf1)

V-B (C2c1)

• Francisella novicida,

• Acidaminococcus sp.

• Lachnospiraceae sp.

• Prevotella sp.

Both subtypes generate staggered DNA breaks distal to the PAM site and enable versatile multiple genomic loci targeting by processing its own guide RNAs.

Cas12 has been engineered as a platform for epigenome editing, and it was recently discovered that Cas12 can indiscriminately chop up single-stranded DNA once activated by a target DNA molecule matching its spacer sequence. This property makes Cas12 a powerful tool for:

− Multiplex genome editing

− Epigenome editing

− Diagnostic of viral diseases

The subtype V-B has a significantly smaller size (1108 aa) than subtype V-A (1307 aa), making it a more suitable tool for gene therapy applications that use a viral delivery system. Furthermore, it is very sensitive to mismatches between the guide RNA and target DNA, thus causing less off-target effects.

Type VI

VI-A

VI-B

VI-C

VI-D

• Leptotrichia buccalis

• Leptotrichia shahii

• Ruminococcus flavefaciens

• Bergeyella zoohelcum

• Prevotella buccae

• Listeria seeligeri

Cas13 targets RNA, not DNA. Once it is activated by a ssRNA sequence complementary to its crRNA spacer, it destroys all nearby RNAs, regardless of their sequence.

Cas13 is an outlier in the CRISPR world because it targets RNA, not DNA. Its applications include:

− Specific RNA knock-down or editing without altering genome sequence

− Diagnostic of viral diseases (RNA viruses)

Subtype VI-B had consistently increased levels of knock-down relative to subtype VI-A. (average of 92.3% versus 40.1%)

crRNA (CRISPR RNA): RNA sequence complementary to the target DNA