Total RNA Extraction and RNA-seq Library Preparation

Total RNA was extracted as previously described (Peng et al., 2021 (link)). Briefly, approximately 0.1 g of symptomatic leaves per sample was ground into a homogenate in liquid nitrogen, and 1 mL of TRIzol (TransGen, Beijing, China) was added to each sample. The total RNA–containing supernatant was washed with an equivalent volume of chloroform; the total RNA was precipitated using isopropanol and dissolved in RNase-free water with RNase inhibitors. The total RNA quality and quantity were assessed using a SpectraMax NanoDrop system (Thermo Fisher Scientific, MA, USA). All samples had RNA integrity numbers higher than 8.0, indicating relatively intact and protein-free RNA. The RNA-seq library was prepared using 4 μg total RNA, with the KAPA Stranded mRNA-Seq Library Preparation Kit (Kapa Biosystems, Roche, Basel, Switzerland). RNA-seq libraries were used for cluster formation with the HiSeq X PE Cluster Kit V2.5 on an Illumina cBOT cluster generation system (Illumina) and sequenced in one lane by using the Illumina HiSeq 2500 platform in the 100 bp paired-end mode.