BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) are widely used in the field of tissue engineering because of their multi-directional differentiation potential. Micro RNAs play an important role in promoting osteogenic differentiation of BMSCs.OBJECTIVE: To investigate the effect of miR-218 and miR-26a on the osteogenic differentiation of rat BMSCs, and to provide reference for the study on osteogenic differentiation of BMSCs and the clinical application. METHODS: The bone marrow of the femur of Wistar rats was extracted and the BMSCs were isolated and cultured to the 3rd generation. MiR-218, miR-26a and polyethyleneimine (PEI) were mixed in a specific ratio to form a miRNA/PEI complex. Meanwhile, a negative control group was established.RESULTS AND CONCLUSION: (1) Rat BMSCs grew well. (2) The optimal concentration of miRNA mimics was 50 nmol/L by MTT method. (3) The expression of alkaline phosphatase and collagen type I mRNA was significantly increased (P < 0.05). (4) Alkaline phosphatase staining showed that compared with the blank control group and the negative control group, the cytoplasm showed obvious coloring. (5) There were a lot of mineralized nodules shown by alizarin red staining. Therefore, miR-218/PEI complex, miR-26a/PEI complex and miR-218+miR-26a/PEI co-transfection complex could promote the osteogenic differentiation of BMSCs. Under the same conditions, the osteogenesis of miR-26a was slightly better than that of miR-218.