Engineer complete resistance to Cotton Leaf Curl Multan virus by the CRISPR/Cas9 system in Nicotiana benthamiana

Clustered, regularly interspaced short palindromic repeats/CRISPR-associated proteins (CRISPR/Cas) based genome editing technology has been developed from the adaptive immune system used by many bacteria and archaea for combating against viruses. Here, we utilize the Cas9 with dual gRNAs designed to target two essential regions of the single-stranded DNA genome of the Cotton Leaf Curl Multan virus (CLCuMuV) which can cleavage the corresponding DNA sequence and thus interrupt viral infection. The transgenic plants expressing Cas9 and dual gRNAs that target different regions of the CLCuMuV genome confer complete resistance to virus infection, thereby demonstrating a novel approach for engineering resistance to geminiviruses.