Extended Data Fig. 5: Phenotype analysis of vsr2VSR3^Cas9VSR4^Cas9-1 mutant.

(a and b) Sequencing analysis of CRISPR-Cas9 edited line of vsr2VSR3^Cas9VSR4^Cas9-1. CRISPR-Cas9 editing of VSR3 (a) and VSR4 (b) was carried out using vsr2 T-DNA insertion mutant as the background. VSR3 and VSR4 share closely related sequences thus with identical gRNA1 and gRNA2 targets. Below each gene model, the nucleotide sequences are shown alongside the corresponding amino acid translations. The nucleotides highlighted in red in the sequences are specific changes from WT sequences. The stop codes are marked with stars. (c and d) Phenotype of 24-day-old plants grown under long-day (c) or short-day (d) growth condition. The experiments were repeated independently three times with similar results. (e and f) Disease symptoms analysis of vsr2VSR3^Cas9VSR4^Cas9-1 mutant. Leaves of WT and vsr2VSR3^Cas9VSR4^Cas9-1 triple mutant after 3 d inoculation with the Pto DC3000 EV (e) or Pto DC3000 expressing avrRpm1 (OD₆₀₀ = 0.001) (f). Loss-of-function mutant of the corresponding resistance gene RPM1 (rpm1) served as an additional control. Scale bars, 1 cm. (g and h) Bacterial growth 3 d after inoculation with Pto DC3000 expressing Pto DC3000 EV (g) or avrRpm1 (h) in the leaves. Each bar represents log₁₀-transformed values of the mean and SD of three biological replicates. Experiments were repeated three times with similar results. cfu, colony-forming units. (i) Trypan blue staining of dead cells in the leaves of WT, vsr2VSR3^Cas9VSR4^Cas9-1, and rpm1 mutants at 12 h after inoculation with Pto DC3000 avrRpm1 (OD₆₀₀ = 0.001). Quantifications of trypan blue staining intensity are shown on the Right. Data are presented as means ± SD of three independent experiments. Scale bars, 5 mm. Different letters above bars in (g-i) indicate a significant difference at the P < 0.05 level by one-way ANOVA with Fisher’s LSD multiple comparisons test. Exact P-values of statistic tests in (h and i) are provided in the Source data file.

Source data