Abstract
Seed aging, a common physiological phenomenon during forage seed storage, is a crucial factor contributing to a loss of vigor, resulting in delayed seed germination and seedling growth, as well as limiting the production of hay. Extensive bodies of research are dedicated to the study of seed aging, with a particular focus on the role of the production and accumulation of reactive oxygen species (ROS) and the ensuing oxidative damage during storage as a primary cause of decreases in seed vigor. To preserve optimal seed vigor, ROS levels must be regulated. The excessive accumulation of ROS can trigger programmed cell death (PCD), which causes the seed to lose vigor permanently. LESION SIMULATING DISEASE (LSD) is one of the proteins that regulate PCD, encodes a small C2C2 zinc finger protein, and plays a molecular function as a transcriptional regulator and scaffold protein. However, genome-wide analysis of LSD genes has not been performed for alfalfa (Medicago sativa), as one of the most important crop species, and, presently, the molecular regulation mechanism of seed aging is not clear enough. Numerous studies have also been unable to explain the essence of seed aging for LSD gene regulating PCD and affecting seed vigor. In this study, we obtained six MsLSD genes in total from the alfalfa (cultivar Zhongmu No. 1) genome. Phylogenetic analysis demonstrated that the MsLSD genes could be classified into three subgroups. In addition, six MsLSD genes were unevenly mapped on three chromosomes in alfalfa. Gene duplication analysis demonstrated that segmental duplication was the key driving force for the expansion of this gene family during evolution. Expression analysis of six MsLSD genes in various tissues and germinating seeds presented their different expressions. RT-qPCR analysis revealed that the expression of three MsLSD genes, including MsLSD2, MsLSD5, and MsLSD6, was significantly induced by seed aging treatment, suggesting that they might play an important role in maintaining seed vigor. Although this finding will provide valuable insights into unveiling the molecular mechanism involved in losing vigor and new strategies to improve alfalfa seed germinability, additional research must comprehensively elucidate the precise pathways through which the MsLSD genes regulate seed vigor.