Comprehensive Identification of the β-Amylase (BAM) Gene Family in Response to Cold Stress in White Clover.

White clover ( L.) is an allopolyploid plant and an excellent perennial legume forage. However, white clover is subjected to various stresses during its growth, with cold stress being one of the major limiting factors affecting its growth and development. Beta-amylase (BAM) is an important starch-hydrolyzing enzyme that plays a significant role in starch degradation and responses to environmental stress. In this study, 21 members of the BAM gene family were identified in the white clover genome. A phylogenetic analysis using from Arabidopsis divided into four groups based on sequence similarity. Through analysis of conserved motifs, gene duplication, synteny analysis, and -acting elements, a deeper understanding of the structure and evolution of in white clover was gained. Additionally, a gene regulatory network (GRN) containing was constructed; gene ontology (GO) annotation analysis revealed close interactions between and AMY (α-amylase) and DPE (4-alpha-glucanotransferase). To determine the function of under various tissues and stresses, RNA-seq datasets were analyzed, showing that most were significantly upregulated in response to biotic and abiotic stresses and the highest expression in leaves. These results were validated through qRT-PCR experiments, indicating their involvement in multiple gene regulatory pathways responding to cold stress. This study provides new insights into the structure, evolution, and function of the white clover BAM gene family, laying the foundation for further exploration of the functional mechanisms through which respond to cold stress.