Metabolomics combined with transcriptomics analysis on ginsenosides accumulation in root of American ginseng plants under foliar applications of brassinolide

Panax quinquefolius L. (American ginseng) is a medicinally important crop with high pharmacological value, but its commercial application is restricted by low ginsenoside yields and insufficient bioavailability. In this study, we performed integrated metabolomic and transcriptomic analyses to explore the regulatory effects of foliar-applied brassinolide (BL) on American ginseng. BL exhibited a dose-dependent effect on the coordination between plant growth and ginsenoside accumulation. Low-concentration BL promoted plant growth and induced the accumulation of five rare ginsenosides (ginsenoside F5, pseudoginsenoside Rt3, majoroside R2, ginsenoside F3, majoroside R1) in roots by 2–21-fold. High-concentration BL displayed no growth-promoting effect but significantly enhanced the accumulation of the same ginsenosides by 11–75-fold. Transcriptomic analysis identified 34 differentially expressed genes (DEGs) involved in the ginsenoside biosynthesis pathway. Weighted gene co-expression network analysis (WGCNA) revealed 16 CYP450/UGT genes and 17 transcription factors as key regulators mediating BL-induced rare ginsenoside biosynthesis. Multi-omics integration further showed that four β-amyrin synthase genes ( PQ0G073990, PQ0G318090, PQ0G569630, novel.5640 ) and two CYP450 genes ( novel.16917, PQ0G686990 ) were co-enriched with oleanane-type saponin Ro O-pentoside and downregulated by high-concentration BL. These results illustrate the dual function of BL in balancing plant growth and secondary metabolism, providing candidate molecular targets for metabolic engineering and sustainable cultivation practices aimed at improving rare ginsenoside production in American ginseng.