Exogenous 24-epibrassinolide enhances chilling tolerance in Phalaenopsis-type Dendrobium through coordinated physiological and transcriptomic responses

Introduction Phalaenopsis-type Dendrobium is a commercially important tropical orchid that is highly sensitive to chilling, which can cause premature leaf senescence and abscission. Methods We evaluated the protective effect of exogenous 24-epibrassinolide (EBR) and integrated time-resolved RNA-seq with physiological traits previously characterized in the same experimental framework. A concentration of 2.0 mg/L was selected for transcriptomic analysis based on the refined dose-response experiment. WGCNA was performed using 18 EBR-treated libraries. Global GO and KEGG outputs were retained as descriptive overviews, whereas quantitative functional inference was based on module-specific analyses with complete multiple-testing-adjusted statistics. Results EBR delayed leaf yellowing and abscission. Differential-expression analysis revealed time-dependent transcriptional responses during chilling and recovery. WGCNA identified trait-associated modules with contrasting temporal relationships to physiological traits: the pink module was positively associated with MDA and proline and negatively associated with chlorophyll, whereas the antiquewhite4 (AW4) module showed the opposite pattern. Pink-module enrichment emphasized central carbon and energy metabolism, lipid turnover, peroxisomal processes, autophagy, and cellular transport/homeostasis. AW4 GO enrichment was dominated by transmembrane and metabolite transport; no AW4 KEGG pathway remained significant after multiple-testing correction. Discussion These results support an association between EBR treatment and coordinated metabolic, membrane-homeostasis, and stress-response adjustments during chilling and recovery in Phalaenopsis-type Dendrobium .