Integration of transcriptomic and metabolomic data reveals the low temperature response gene regulatory network of germination in soybean (Glycine max L.)

Low-temperature stress (<6 ℃) during soybean germination significantly delays germination timing, reduces germination percentage, and ultimately decreases yield. To enhance stress-resilient yield stability, we identified low-temperature-tolerant germplasm and characterized associated genetic resources. Using extreme cold-phenotype accessions (chilling-tolerant C143 and chilling-sensitive K2C037) during the germination process at 6 ℃, we investigated cold-response mechanisms through transcriptomic and metabolomic analyses of both imbibition and germination stages. Our analyses revealed 437 differentially expressed genes (DEGs) and 28 differentially accumulated metabolites (DAMs) in tolerant accessions, compared to 152 DEGs and 38 DAMs in sensitive accessions post-imbibition phase. Post-germination phases, we identified 232 DEGs and 45 DAMs in tolerant accessions versus 209 DEGs and 22 DAMs in sensitive accessions. Integration of Pearson correlation analysis between DEGs and DAMs with weighted gene co-expression network analysis (WGCNA) identified 16 and 17 candidate cold-response genes for imbibition and germination stages, respectively. In the post-imbibition phase, sixteen low-temperature-responsive candidate genes were identified, including LTI65, CAT2, PEX19–1, RVE1, BGLU44, and ECT2. These genes showed co-expression with known stress-related genes and were functionally enriched in plant hormone signaling, cold response, and fatty acid metabolism pathways. After germination, key candidate genes included transcription factors (YABBY7, YABBY13, YABBY16, ATHB-16), transporters (SWEET4, SWEET4-like, SULTR2;1), and metabolic genes (APL3, LTP3), whose co-expression networks were primarily associated with plant hormone responses, peroxisome function, fatty acid metabolism, and cold stress response. These findings provide novel genetic targets for improving soybean cold tolerance during critical early growth stages, offering potential strategies for breeding chilling-tolerant soybean varieties.