Integrated transcriptomic and metabolomic analyses of Caucasian clover (Trifolium ambiguum Bieb.) in response to freezing stress

Caucasian clover (Trifolium ambiguum Bieb.) is a perennial leguminous herbage with unique properties, such as resistance to cold, drought, and grazing. However, the response of Caucasian clover to freezing stress based on gene expression level and metabolite accumulation is unknown. This study performed transcriptomic and metabolomic analyses to investigate the response mechanism of Caucasian clover under freezing stress. Transcriptome analysis detected 6792 differentially expressed genes (DEGs) in Caucasian clover under freezing stress. These DEGs were significantly enriched in carbohydrate and energy metabolism, lipid metabolism, and biosynthesis of secondary metabolites. Meanwhile, 139 differentially accumulated metabolites (DAMs) were identified in Caucasian clover under freezing stress. These DAMs were mainly enriched in amino acid metabolism and biosynthesis of secondary metabolites. Furthermore, the gene-to-metabolite network was used to determine the genes and metabolites involved in polyphenol biosynthesis and osmoprotectants metabolism. Therefore, this study provides insights into the complex regulatory strategy of Caucasian clover response to freezing stress.