EE-Induced SAR

SAR assay was performed as described previously (Hilfiker et al., 2014). Pseudomonas syringae pv. tomato DC3000 (Pst) was grown in King’s B medium containing 50 μg/ml rifampicin at 28°C. Overnight log phase cultures were washes three times with 10 mM MgCl₂ and diluted to OD₆₀₀ of 0.0005 for leaf inoculation. To induce SAR, three fully developped leaves of each of six Col-0 and lecrk-I.8 plants were treated with 2 μl × 2 μl of EE from the abaxial side of the leaf. Five days after the treatment, EE was carefully removed with a brush and three untreated leaves distal to the site of EE treatment were inoculated with a suspension of Pst at OD₆₀₀ 0.0005 in 10 mM MgCl₂ from the abaxial side with a 1 ml needleless syringe. The same amount of untreated plants was inoculated with Pst and served as controls. Growth of Pst in inoculated leaves was measured 48 h later by serial dilutions on LB plates.