RNA-Seq Analysis of Tender Roots

The tender roots at the bud stage under two treatments for 12 h were used as samples for RNA-Seq analysis, and each treatment had three biological replicates. These samples were rapidly frozen in liquid nitrogen. And then RNA was extracted by RNApure Plant Kit (CW0559, CWBIO, Beijing, China) following the manufacturer’s instructions. The RNA was measured in 1% agarose gel electrophoresis and NanoDrop instrumentation (OneC, Thermo, Waltham, MA, USA) using the methods of Zhang [20 (link)], which determined whether the quality of the RNA was qualified. The qualified RNA of these samples underwent RNA-Seq by Bio-maker (Beijing, China), while those with no reference genome were analyzed by Biocloud (https://international.biocloud.net/, accessed on 1 January 2022). UniGene was analyzed by Trinity software [32 (link)], while the differentially expressed genes (DEGs) were assessed by DESeq2 software [33 (link)]. In addition, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases were used for gene and pathway enrichment analyses [34 (link)].