Rhododendron liliiflorum H. Lév., with white outer edges and yellow inner edges of petals, is an ornamental flower that originated in China. In this study, we analysed the white (W) and yellow (Y) parts of R. liliiflorum flowers by RNA sequencing. Then, unigene assembly, unigene annotation, and classification of Eukaryotic Orthologous Groups (KOGs) were performed. Gene ontology (GO) classification and pathway enrichment analysis for unigenes were also conducted. A total of 219,221 transcripts and 180,677 unigenes of R. liliiflorum were obtained from 48.52 Gb of clean reads. Differentially expressed gene (DEG) analysis indicated that 2310 unigenes were upregulated and 3062 were downregulated in W vs. Y. Thirty-six of these DEGs were involved in the flavonoid biosynthesis pathway. Pathway enrichment analysis showed that DEGs were significantly enriched in phenylpropanoid, flavonoid, and isoflavone biosynthesis. The expression of dihydroflavonol-4-reductase (DFR) and chalcone synthase (CHS) may affect differences in R. liliiflorum flower colour. The findings on flavonoid biosynthesis and other related genes in this study will provide guidance for exploring the mechanism of flower colour formation in Rhododendron.

Among ornamental plants, the colour of the petals is an important feature. However, the reason for the colour differences of Rhododendron liliiflorum remains unclear. To reveal the differences in the colour of R. liliiflorum, high-efficiency liquid chromatographic collar (UPLC–MS/MS) technology was used to study the yellow and white parts of R. liliiflorum. A total of 1187 metabolites were identified in R. liliiflorum petals, including 339 flavonoid metabolites. Seventy-eight types of flavonoids in these metabolites were found in the yellow and white parts of R. liliiflorum petals, along with 11 other significantly enriched substances. Combining gene expression-related data with differential metabolite data demonstrated effects of enrichment in the flavanonols (fustin), flavonols (epiafzelechin and afzelechin), and flavanones (pinocembrin) of flavonoid biosynthesis; glyccitin, 6″-O-malonylgenistin, and 6-hydroxydaidzein of isoflavonoid biosynthesis; and anthocyanin biosynthesis of malvidin-3-O-galactoside (primulin), delphinidin-3-O-rutinoside, cyanidin-3-O-glucoside (kuromanin), and cyanidin-3-O-rutinoside (keracyanin), which are potentially the contributing factors responsible for the differences in petal colour in R. liliiflorum. This study establishes a connection between the differential metabolites underlying the color differences in the petals of R. liliiflorum and the gene expression in R. liliiflorum. This will provide a foundation for subsequent research on the regulation of flower color in R. liliiflorum and have profound implications for horticultural applications of R. liliiflorum.