Halogenated acid dehalogenases (HADs) hydrolyze a variety of organic phosphate substrates. They have been shown to play important roles in various physiological processes (e.g. phosphorus metabolism and stress tolerance) by few studies. In this study, 24 and 29 HAD proteins were identified in Medicago truncatula and M. sativa, respectively. Conserved motif analysis showed that most HAD proteins contain acyltransferases and phosphatase domains, and all HAD proteins have a common conserved motif (DXD). Gene duplication analysis showed that tandem duplication is the main driver of HAD gene evolution with intense purification pressure. Moreover, cis-acting element analysis for promoter region implied that HAD gene expression in Medicago is closely related to phytohormones, light, and various stresses. Available transcription data revealed that specific HAD genes were highly expressed in leaf, flower and mature seed. HAD genes responding to phosphorus deficiency, low temperature, high temperature, dehydration and salt stress were found. Meanwhile, members of HADs that may be involved in nodule organogenesis and symbiosis with arbuscular mycorrhizal fungi were identified. These results provide a basis for further functional studies of the HAD gene family and potential applications for genetic improvement in the context of stress and plant-microbe interactions.