CRISPR Plasmids: Cascade-Cas3

Cas3 is a unique member of the CRISPR family. After nicking the target DNA, Cas3 initiates single-strand DNA degradation. This degradation can continue for many kilobases in one or both directions from the target region, effectively turning Cas3 into a DNA “shredder” system that can cause large genomic deletions. Compared to deletions generated by Cas9, large deletions created using Cas3 have fewer off-target alterations and require only a single gRNA.

Cas3 must be paired with the Cascade (CRISPR-associated complex for antiviral defense) complex, comprised of a combination of Cas5, Cas6, Cas7, Cas8, and Cas11. Cascade is directed to target DNA using a gRNA and recruits Cas3. After nicking the non-targeted strand, Cas3 starts unraveling DNA using its helicase properties, cleaving the targeted strand non-specifically along the way. Cas3 initiates a final double-strand break, either due to a boundary defined by anti-CRISPR proteins or running into other proteins bound to DNA, and the break can be repaired by the cell’s endogenous repair mechanisms.

Cas3 belongs to the Class 1 family of CRISPR systems, the most abundant type found in bacteria and archaea. While abundant in nature, Class 1 systems are largely underutilized compared to their Class 2 counterparts like Cas9. This is attributed in part to the difficulties in getting all components of the Cascade complex to express equally. To combat this, some CRISPR-Cas3 plasmids take an all-in-one approach and include Cas3 and Cascade on one single plasmid, though many still separate the individual Cas components.

Figure 1: Overview of Cascade-Cas3 mechanism. Created with BioRender.com.

Browse, sort, or search the tables below for CRISPR Cascade-Cas3 plasmids.
Plasmids are available for expression in mammalian systems, bacteria, and plants.

Mammalian

Bacteria

Plant

Last reviewed on: January 30, 2025