Phenylalanine ammonia lyase (PAL) is the first enzyme in the phenylpropanoid pathway and plays a critical role in plant growth, development and stress defence. However, there have been few reports of the PAL gene family in lucerne (also known as alfalfa, Medicago sativa L.), one of the most important forage legume species worldwide. In this study, we report that PAL in lucerne is encoded by a family of seven genes: MsPAL1-MsPAL7. Furthermore, a comprehensive genome-wide bioinformatics analysis of the MsPAL gene family is presented, including chromosomal locations, phylogenetic relationships, gene structures and conserved motifs. The cis-elements and potential biological functions of these genes were investigated, revealing the potential roles of MsPAL members in response to various stresses. RT-qPCR results showed that the expression of MsPAL6 was significantly upregulated under both salinity- and waterlogging-stress conditions. Other MsPAL members such as MsPAL1 and MsPAL2 were downregulated under saline conditions and upregulated significantly after waterlogging stress. Our findings provide useful information for further practical analyses and for the genetic improvement of abiotic stress tolerance of lucerne.