Table 5.
Comparison of different CRISPR/Cas and Fanzor protein properties.
| CRISPR/Cas9 | CRISPR/Cas12 | CRISPR/Cas13 | CRISPR/Cas14 | Fanzor Proteins | |
|---|---|---|---|---|---|
| DNA catalytic domain | RuvC, HNH | RuvC-like nuclease domain, Nuc-domain | HEPN domains | RuvC | RuvC-like nuclease domain |
| Target | Double-stranded DNA | Double-stranded DNA | Single-stranded RNA | Single-stranded DNA | Double-stranded DNA |
| Collateral activity | No | Yes | Yes | Yes | No |
| DNA recognition | sgRNA (crRNA in complex with tracrRNA) | crRNA | crRNA | crRNA and tracrRNA | fRNA or ωRNA |
| PAM requirements | NGG, NAG for SpCas9, and other PAM variants for Cas9 orthologs (for details, see Table S4) | TTTN, TTTV (V = G, C, or A) For AsCpf1 from Acidaminococcus or LbCpf1 from Lachnospiraceae, and other PAM variants for Cas12 orthologs [362,363,364,365,366,367,368] |
Requires protospacer flanking sequence–A, U, or C | None | Target adjacent motif preference is diverse, with a GC preference observed for the viral Fanzor proteins and AT preferences for the eukaryotic Fanzor proteins |
| Specificity | Regular SpCas9 tolerates mismatches, but high-fidelity variants exist | Cas12a has been successfully used for gene editing in vivo without any deleterious off-target effects | RNA-editor, no damage to DNA may occur | Cleaves ssDNA with high fidelity-sensitive to even a single mismatch in the target sequence | Needs further investigations |
| Ease of delivery | Easily delivered using multiple techniques | Easily delivered using multiple techniques | Easily delivered using multiple techniques | Easily delivered using multiple techniques | Needs further investigations |
| Limitations | GC-rich DNA targets Possible off-target effects (especially for SpCas9) |
AT-rich DNA targets | Needs to be constitutively expressed to maintain the editing effect | Targets ssDNA | Unknown |
| Multiplexing | Easy | Easy | Easy | Easy | Possible |
| Active clinical trials | Yes (64/130) |
Yes (2/130) |
No | No | No |
| Use in academic laboratories | 26,717 results in PubMed® (https://pubmed.ncbi.nlm.nih.gov/, accessed on 2 November 2023 with search term “CRISPR Cas9” | 1684 results in PubMed® (https://pubmed.ncbi.nlm.nih.gov/, accessed on 2 November 2023 with search term “CRISPR Cas12a or Cpf1” | 373 results in PubMed® (https://pubmed.ncbi.nlm.nih.gov/, accessed on 2 November 2023 with search term “CRISPR Cas13” | 28 results in PubMed® (https://pubmed.ncbi.nlm.nih.gov/, accessed on 2 November 2023 with search term “CRISPR Cas14” | 4 results in PubMed® (https://pubmed.ncbi.nlm.nih.gov/, accessed on 2-November 2023 with search term “Fanzor” |
| First time mentioned | 2011 | 2017 | 2017 | 2018 | 2013, 2023 |