Targeted large fragment deletion in plants using paired crRNAs with type I CRISPR system

Abstract

SummaryThe CRISPR‐Cas systems have been widely used as genome editing tools, with type II and V systems typically introducing small indels, and type I system mediating long‐range deletions. However, the precision of type I systems for large fragment deletion is still remained to be optimized. Here, we developed a compact Cascade‐Cas3 Dvu I‐C system with Cas11c for plant genome editing. The Dvu I‐C system was efficient to introduce controllable large fragment deletion up to at least 20 kb using paired crRNAs. The paired‐crRNAs design also improved the controllability of deletions for the type I‐E system. Dvu I‐C system was sensitive to spacer length and mismatch, which was benefit for target specificity. In addition, we showed that the Dvu I‐C system was efficient for generating stable transgenic lines in maize and rice with the editing efficiency up to 86.67%. Overall, Dvu I‐C system we developed here is powerful for achieving controllable large fragment deletions.