2.5 RNA preparation and construction of nanopore sequencing library
Total RNA was isolated using the FastPure Universal Plant Total RNA Isolation Kit (Vazyme, Nanjing, China) according to the manufacturer’s instructions. RNA was cleaned and concentrated by NEB Next Poly(A) mRNA Magnetic Isolation Module (E7490S) according to the manufacturer’s instructions. Each nanopore DRS and transcriptome sequencing sample contained 20 μg total RNA. Prepared RNA of each sample was used for a nanopore DRS library preparation using the Oxford Nanopore DRS protocol (SQK-RNA002, Oxford Nanopore Technologies) (Yahara et al., 2021). This work was done in collaboration with Wuhan Benagen Technology Co, Ltd, China.