2.2 RNA extraction and transcriptome sequencing
Total RNA of all test samples was isolated using the
Quick-RNATM MicroPrep Kit (ZYMO Research, USA)
according to the manufacturer’s protocol. The integrity of the total RNA
was analyzed using 1.5 % agarose gel electrophoresis (Cui et al.,
2017). The quality and concentration were analyzed on NanoDrop 2000
spectrophotometer (Thermo Scientific, USA). The cDNA was synthesized by
following the instructions from RTTM All-in-One Master
Mix Kit (Herogen Biotech, USA). Transcriptome sequencing was performed
at Novogen Co., Ltd. Beijing, China, and the samples were sequenced on
the Illumina Hiseq 2500 platform. The raw reads were curated by removing
adaptor sequences and reads of low quality, then assembled into unigenes
using Trinity (Huang et al., 2018; Grabherr et al., 2011).