The prolyl hydroxylase inhibitor dimethyloxallyl glycine (DMOG) has been increasingly studied with regards to stem cell therapy. Previous studies have demonstrated that endogenous mesenchymal stem cells (MSCs) may be mobilized into peripheral circulation by pharmaceutical preconditioning. In addition, our previous study confirmed that DMOG, as a novel mobilization agent, could induce mouse/rat MSC migration into peripheral blood circulation. Therefore, the present study conducted studies to characterize bone marrow‑derived MSCs (BM‑MSCs) collected from mice following DMOG intraperitoneal injection. The surface antigen immune phenotype, differentiation capability, proliferative ability, migratory capacity and paracrine capacity of the BM‑MSCs collected from DMOG‑preconditioned mice (DBM‑MSCs) or normal saline‑treated mice (NBM‑MSCs) were evaluated by means of flow cytometry, differentiation induction, Cell Counting kit‑8, Transwell assay and enzyme-linked immunosorbent assay, respectively. Compared with NBM‑MSCs, DBM-MSCs displayed a similar immune phenotype and multilineage differentiation capability, reduced proliferative ability and migratory capacity, and similar transforming growth factor and platelet-derived growth factor secretion capacity. These results provide a novel insight into the biological properties of BM‑MSCs from mice preconditioned with DMOG. DBM-MSCs exhibited slightly distinct characteristics to NBM-MSCs; however, they may have therapeutic potential for future stem cell therapy. In addition, the present study suggested that DMOG may be used as a novel mobilization agent in future clinical trials as no adverse effects were observed.