RNA Isolation and Sequencing of Rice Seedlings

The methods used for RNA isolation are described in Li et al. (2016) [16 (link)]. Two biological replicates were used for each genotype. The total RNA of shoots and roots was extracted from Dongdao-4 and Jigeng-88 seedlings using RNAiso reagent (TaKaRa Biotechnology Co., Ltd., Kyoto, Japan). RNA integrity and quality were assayed with agarose gel electrophoresis and a Bioanalyzer using an Agilent RNA 6000 Nano Chip (Agilent Technologies, Santa Clara, CA, USA). Samples with RNA integrity number values >8 were used for reverse- transcription into first-strand cDNA. cDNA libraries were prepared according to the manufacturer’s protocol (Illumina, San Diego, CA, USA) and assayed using the BioAnalyzer 2100 system and qPCR (Kapa Biosystems, Woburn, MA, USA). Subsequently, the libraries were sequenced on an Illumina HiSeq 2000 instrument.