Rna reagent kit

Total RNA was extracted from the TW, OW and NW using an RNA reagent kit (DP441; Tiangen Biotech, Beijing, China) according to the manufacturer’s protocol. Nine transcriptome libraries were constructed for 150-bp paired-end Illumina high-throughput sequencing of mRNAs and lncRNAs using the Illumina HiSeq™ 4000 platform (Illumina, USA) by Gene Denovo Biotechnology Co. (Guangzhou, China). The post sequencing data filtering and analysis methodology have been described in a previous study [57 (link)]. Cufflinks and Cuffcompare software [58 (link), 59 (link)] were used to assemble transcripts and compare sequences to known sequences in the C. bungei genome (unpublished). The fragments per kilobase of transcript per million mapped reads (FPKM) value was used as an indicator to evaluate gene expression [59 (link)]. In the present study, transcripts with a fold change of ≥1.5 or ≤ − 1.5 and a P value of < 0.05 between various comparison groups were considered significant DEGs.

Total RNA of TW, OW and NW was extracted using an RNA reagent kit (DP441; Tiangen Biotech, Beijing, China) according to the manufacturer’s protocol. Three replicates were included for each type of wood. After total RNA was extracted, rRNAs were removed to retain mRNAs. Then, nine transcriptome libraries were constructed for mRNA and small RNA Illumina high-throughput sequencing. The libraries were sequenced using an Illumina HiSeqTM 4000 platform (Illumina, San Diego, CA, USA) by Gene Denovo Biotechnology Co. (Guangzhou, China).
After sequencing, the reads were further filtered to obtain high-quality clean reads according to the following rules: (1) reads containing adapters were removed; (2) reads consisting of all A bases were removed; (3) reads containing more than 10% of unknown nucleotides (N) were removed; and (4) low-quality reads containing more than 50% low-quality (Q-value ≤ 20) bases were removed. rRNA was removed from the reads of each sample, and then the reads were mapped to the reference genome by TopHat2.