Enhancing tolerance to Phytophthora spp. in eggplant through DMR6–1 CRISPR/Cas9 knockout

Abstract

Agricultural production is affected by the worsening effects of climate change with severe yield losses caused by rising temperatures, water scarcity and consequent modifications in the interactions between crops, pests, and pathogens. The availability of stress-tolerant plants will thus be a key point to guarantee the world food security in the next future. To this purpose, a significant contribution might be provided by the New Genomic Techniques (NGTs), such as CRISPR/Cas9, which allow to insert targeted modifications in the plants’ genomes offering new opportunities for crop improvement. Susceptibility genes encode proteins that pathogens can take advantage of during the colonization process, and their disabling confers a broad-spectrum and long-lasting pathogen tolerance to the plant. Among the S-genes, Downy Mildew Resistance 6 (DMR6) encodes an enzyme involved in Salicylic Acid (SA) degradation, and its inactivation in other Solanaceae species has proven to increase SA levels and confer tolerance to a broad spectrum of pathogens. We identified two orthologs of this gene in eggplant's genome, namely SmDMR6–1 and SmDMR6–2. In the ‘Black Beauty’ cultivar, only SmDMR6–1 expression significantly increased upon infection by the two oomycetes Phytophthora infestans and Phytophthora capsici, suggesting its involvement in the regulation of plant responses to biotic stresses. Here we report, for the first time in eggplant, the knockout of SmDMR6–1 gene through CRISPR/Cas9 technology. The regenerated T₀ plants were screened by Sanger sequencing and one was selected and self-pollinated to generate T₁ and then T₂ plants. The mutant lines were subjected to pathogen assays which highlighted an increased tolerance to infection by P. infestans and P. capsici, if compared to non-edited plants.