RPS5A Promoter-Driven Cas9 Produces Heritable Virus-Induced Genome Editing in .

The virus-induced genome editing (VIGE) system aims to induce targeted mutations in seeds without requiring any tissue culture. Here, we show that tobacco rattle virus (TRV) harboring guide RNA (gRNA) edits germ cells in a wild tobacco, , that expresses Cas9 (SpCas9). We first generated transgenic plants expressing SpCas9 under the control of 35S promoter and infected rosette leaves with TRV carrying gRNA. Gene-edited seeds were not found in the progeny of the infected . Next, the ribosomal protein S5 A (RPS5A) promoter fused to SpCas9 was employed to induce the heritable gene editing with TRV. The RPS5A promoter-driven SpCas9 successfully produced monoallelic mutations at three target genes in seeds with TRV-delivered guide RNA. These monoallelic mutations were found in 2%-6% seeds among M progenies. This editing method provides an alternative way to increase the heritable editing efficacy of VIGE.