Conferring DNA virus resistance with high specificity in plants using virus-inducible genome-editing system

Xiang Ji,Feng Zhang,Yi Zhang,Huawei Zhang,Caixia Gao,Xiaomin Si

doi:10.1186/s13059-018-1580-4

Abstract

The CRISPR/Cas9 system has recently been engineered to confer resistance to geminiviruses in plants. However, we show here that the usefulness of this antiviral strategy is undermined by off-target effects identified by deep sequencing in Arabidopsis. We construct two virus-inducible CRISPR/Cas9 vectors that efficiently inhibit beet severe curly top virus (BSCTV) accumulation in both transient assays (Nicotiana benthamiana) and transgenic lines (Arabidopsis). Deep sequencing detects no off-target effect in candidate sites of the transgenic Arabidopsis. This kind of virus-inducible genome-editing system should be widely applicable for generating virus-resistant plants without off-target costs.

Highlights

Agriculture worldwide is threatened by plant pathogens, such as plant viruses, which account for major losses in crop yields and revenues
The CRISPR/ Cas9 system originated in bacteria and archaea as an adaptive immune system and has been engineered to be a versatile genome-editing tool with applications in numerous organisms including targeting the genome of geminiviruses to inhibit their multiplication in plants
We searched for potential off-target sites of the C3 sgRNA sequence with Cas-OFFinder software [17] using either the canonical NGG or non-canonical NAG protospacer-associated motifs (PAMs) for SpCas9 in Arabidopsis and found that the minimal mismatches were 3 bp

Summary

Introduction

Agriculture worldwide is threatened by plant pathogens, such as plant viruses, which account for major losses in crop yields and revenues. It has been highly desirable to develop plant varieties with improved geminiviruses resistance [1, 2]. Limited success was achieved through conventional transgenic approaches such as the use of pathogen-derived resistance (PDR) and RNA interference (RNAi) [3]. Recent researches using the CRISPR/Cas9-mediated genome-editing technology showed promise to significantly reduce or even abolish disease symptoms in plants [4,5,6,7,8]. The CRISPR/ Cas system originated in bacteria and archaea as an adaptive immune system and has been engineered to be a versatile genome-editing tool with applications in numerous organisms including targeting the genome of geminiviruses to inhibit their multiplication in plants.

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Conclusion