Salicylic Acid Extraction and Quantification

The SA extraction was performed as described by Sun et al. (2014) (link). In brief, 100–200 mg root samples were ground to powder in liquid nitrogen and homogenised twice with the cold extraction buffer (methanol:ddH₂O:glacial acetic acid = 80:19:1) for oscillation overnight in the dark at 4°C. The extraction was evaporated dry with N₂. The dry powder was dissolved with 300 μL methanols; and the solution was filtered by 0.22 μm filters. SA content was measured by HPLC-MS/MS performed by AB SCIEX QTRAP 4500 system (AB SCIEX, Foster City, CA, United States) as described by Zhou et al. (2018) (link). The experiment was repeated three times, with samples containing more than three seedlings.