show Abstracthide AbstractLeaves are the most critical portion of forage crops such as alfalfa (Medicago sativa). Leaf senescence caused by environmental stresses significantly impacts the biomass and quality of forages. To understand the molecular mechanisms and identify the key regulator of the salt stress-induced leaf senescence process, we conducted a simple and effective salt stress-induced leaf senescence assay in Medicago truncatula, which was followed by RNA-Seq analysis and physiological and biochemical characterization. By comparing the observed expression data with that derived from dark-induced leaf senescence at different time points, we identified 3001, 3787, and 4419 senescence-associated genes (SAGs) for salt stress-induced leaf senescence at 2, 4, and 6 days, respectively. There were 1546 SAGs shared by darkness and salt stress treatment across the three time points. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses showed that the 1546 SAGs were mainly related to protein and amino acids metabolism, photosynthesis, chlorophyll metabolism, and hormone signaling during leaf senescence. These results clarify SAGs under salt stress in M. truncatula and provide new insights and additional genetic resources for further forage crop breeding.