Targeted mutation in eIF4G gene in rice

Rice tungro disease (RTD), caused by the interaction between rice tungro spherical virus (RTSV) and rice tungro bacilliform virus (RTBV) is a serious constraint in rice production. Natural RTSV resistance is due to a mutation in the translation initiation factor 4 gamma (eIF4G) gene. To develop novel alleles for resistance to RTD through CRISPR/Cas9 mediated genome editing, a guide RNA sequence, having higher GC content and fewer off -targets, targeting exon 7 of eIF4G was designed and cloned into a binary vector, pRGEB32. This construct was used for generating mutations in rice cultivar ASD16 through Agrobacterium-mediated transformation. A target region in the thirteen putative mutants generated in this experiment was sequenced, of which six were found to possess indels. The analysis of the predicted amino acid sequence identified deletion mutants that possess near full -length eIF4G gene which can impart resistance to RTSV.