Strains, Culture Conditions, PCR, and Statistical Test

Pyricularia oryzae strains (Table ​Table11) were stored on filter disks at -20°C. Growth tests of P. oryzae were performed in complete medium (CM) (Talbot et al., 1993), minimal medium (MM) (CM medium without peptone, yeast extract, and casamino acid), CM or MM media in which 1% (w/v) glucose was replaced by 1% (v/v) glycerol, 50 mM sodium acetate, 5 mM sodium pyruvate, 1.15% sodium glutamate, 1% glutamine, or 1% olive oil, and CM or MM media supplemented with different chemicals (0.8 M NaCl, 1 M sorbitol, 0.5 mM H₂O₂ or 0.8 mM Paraquat) at 25°C under a light–dark cycle (16 h–8 h) or under continuous dark. Fungal samples were ground in liquid nitrogen and total RNA was extracted with the Trizol method following the manufacturer’s procedure (TaKaRa, Japan). Total RNA (500 ng) was reverse transcribed into first-strand cDNA using a PrimeScript^TM RT reagent kit with gDNA Eraser (Takara, Japan). Quantitative real time PCR (qPCR) was conducted with five biological replicates using SYBR Premix Ex Taq (Tli RNaseH Plus) kit following the manufacturer’s protocol (TaKaRa, Japan) on a Real-Time PCR Detection System Mastercycler (Eppendorf, Germany). To compare relative abundance of transcripts, average threshold cycle (CT) was normalized to β-TUBULIN and H3 for each strain as 2^-ΔCT, where ΔCT = [CT_gene – (CT_β-TUBULIN + CT_H3)/2]. Fold changes among different strains were calculated as 2^-ΔΔCT, where ΔΔCT = ΔCT_{strain_1}-ΔCT_{strain_2} (Livak and Schmittgen, 2001). The PCR primers used in this study are listed in Supplementary Table S1. Tukey’s HSD test was used for all experimental data in this study (Tang and Zhang, 2013).

Pyricularia oryzae strains used in this study.