We have isolated a cardiomyogenic (CMG) cell line from murine bone marrow stroma. Stromal cells were immortalized, treated with 5‐azacytidine, and spontaneous beating cells were repeatedly screened for. The cells showed a fibroblast‐like morphology. However, this morphology changed after 5‐azacytidine treatment in about 30% of the cells, which connected with adjoining cells after 1 week, formed myotube‐like structures and began spontaneous beating after 2 weeks, and beat synchronously after 3 weeks. These cells expressed atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP). Electron microscopy revealed a cardiomyocyte‐like ultrastructure including typical sarcomeres and atrial granules. They had sinus node‐like or ventricular cell‐like action potentials. Analysis of the isoform of contractile protein genes, such as myosin and α‐actin, indicated that their phenotype was similar to fetal ventricular cardiomyocytes. These cells expressed Nkx2.5, GATA4, TEF‐1, and MEF2‐C mRNA before 5‐azacytidine treatment, and expressed MEF2‐A and MEF2‐D after treatment. This new cell line provides a powerful model for the study of cardiomyocyte transplantation.