Genome Editing of Indica Rice ASD16 for Imparting Resistance Against Rice Tungro Disease

Rice tungro disease (RTD) is caused by the joint infection of rice tungro bacilliform virus and rice tungro spherical virus (RTSV) and is the most important viral disease that limits rice production and productivity in major rice growing regions of South and Southeast Asia. Cultivars with RTSV resistance alone could significantly reduce RTD incidence in the fields. RTSV resistance is conferred by in-frame mutations in SVLFPNLAGKSYVV motif (mainly NL and YV residue) in translation initiation factor 4 gamma (eIF4G) gene. Here, we report CRISPR/Cas9-mediated genome editing to create novel in-frame mutations targeting this residue in indica rice. An average mutation efficiency of 46.19% was observed and 30 T0 independent events had in-frame mutations in one of the alleles of eIF4G gene. Twenty events with homozygous in-frame mutations were identified in T1 generation. These mutations were faithfully inherited to T2 generation. Bioassay on transgene-free homozygous T2 mutants suggested that mutations specifically affecting the target residue could successfully confer resistance to RTD. These homozygous mutants did not have any unintended off-target editing and were agronomically similar in performance to their wild type counterpart.