CRISPR/Cas9 targeted mutagenesis of the susceptibility gene PMR4 in tomato ‘San Marzano’ and ‘Oxheart’
Li R.1, Maioli A.1, Zhe Y.2, Bai Y.2, Milani AM.1, Valentino D.1, Comino C.1, Lanteri S.1, Moglia A.1, Acquadro A.1
- Abstract
- Late blight (LB), caused by Phytophthora infestans, and powdery mildew (PM), caused by species within the Erysiphaceae family, are the most destructive and widespread diseases in tomato (Solanum lycopersicum L.). The control of these pathogens extensively relies on fungicide application, leading to economic losses and environment impact. The disabling of susceptibility (S) genes, which facilitate pathogen infection, may provide a broad-spectrum and durable type of resistance. To this purpose, the CRISPR/Cas9 gene editing represents the most suitable and fast strategy. We selected the Powdery Mildew Resistant 4 (PMR4) gene, whose disabling has been shown to confer PM resistance in Arabidopsis and in the tomato variety ‘Moneymaker’. In order to verify whether the effect of disabling S genes is background-dependent, we knocked-out the PMR4 gene in two widely cultivated varieties, ‘San Marzano’ and ‘Oxheart’. CRISPR mutants were generated by using 4 sgRNAs targeting the FKS1dom1 and Glucan-synthase domain of the PMR4 protein in order to maximize editing efficiency. In all, 87 tomato pmr4 transformants were obtained (T0 generation) of which 70 in ‘San Marzano’ and 17 in ‘Oxheart’. Twelve of ‘San Marzano’ mutants were tested and resulted positive for the presence of Cas9 gene. The Sanger analysis performed on target editing regions revealed 3 edited plants with large deletions (~2kb) and 9 showing a prevalence of 7bp deletions. In the next future, mutants will be test with PM and LB to quantify the level of resistance.
- Presented by
- Ruiling Li
- Institution
- 1 Department of Agricultural, Forest and Food Science, University of Turin, Italy 2 Plant Breeding, Wageningen University & research, Droevendaalsesteeg 1, 6708PB Wageningen, The Netherlands




































































































