Cas13a-based multiplex RNA targeting for potato virus Y

Abstract

AbstractCRISPR-Cas systems endow the bacterial and archaeal species with adaptive immunity mechanisms to fend off invading phages and foreign plasmids. The class 2 type VI CRISPR/Cas effector Cas13a has been harnessed to confer the protection against RNA viruses in diverse eukaryotic species. However, whether gRNA targeting sites have effects on the efficiency of RNA viruses inhibition is unknown. Here we repurpose of CRISPR/Cas13a coupled with an endogenous tRNA-processing system (polycistronic tRNA-gRNA, PTG) to target four genes of potato virus Y (PVY). Expression of Cas13a and four different gRNAs were evidenced in transgenic potato lines expressing Cas13a/PTG construct. We show that the multiple gRNAs transgenic plants could suppress PVY accumulation and disease symptoms at a similar level to transgenic plants expressing single gRNA. Collectively, our study suggests that the Cas13-based multiplex RNA targeting system can be used to engineer resistances to RNA viruses in plants, and the number of gRNAs target sites have no effect on CRISPR/Cas13a-mediated viral interference in plants.