Identification of the Plant Defensin (MsPDF) Gene Family in Medicago sativa and Analysis of Expression Patterns Under Abiotic Stress

<i>Medicago sativa</i> L. (alfalfa) is a major forage crop due to its high yield and stress resilience. However, its growth and productivity are often compromised by abiotic stresses, including cold, drought, and salinity. The plant defensin (PDF) gene family plays a crucial role in resistance to abiotic stress. In this study, a total of 11 MsPDF gene family members were identified in the alfalfa genome and classified into three groups. Phylogenetic and conserved motif analyses revealed that the MsPDF genes are highly conserved. Promoter analysis, gene regulatory network analysis (GRN), and gene ontology (GO)-enrichment analyses were used to infer the potential functions of <i>MsPDF</i> genes. The results showed that the gene actively responds to abiotic stress, participates in phytohormonal responses, and regulates plant growth and development through gene interactions. Transcriptome and qRT-PCR analyses showed that most of the <i>MsPDF</i> genes were significantly up-regulated under cold, drought, and salinity stresses. Among them, the <i>MsPDF03</i> exhibited superior performance under cold stress. The <i>MsPDF04</i>, <i>MsPDF08</i>, and <i>MsPDF09</i> genes were able to respond positively to drought and salt stresses. Finally, the monomeric, dimeric, and tetrameric structures of the proteins encoded by the <i>MsPDF</i> genes were predicted using AlphaFold 2 software. This study lays the foundation for the identification and evolutionary relationship analysis of the MsPDF gene family, and provides a new reference for subsequent research on abiotic stress resistance.