Sazacytidine treatment of mouse C3HlOT1/2 embryonic fibroblasts converts them to myoblasts at a frequency suggesting alteration of one or only a few closely linked regulatory loci. Assuming such loci to be differentially expressed as poly (A)+ RNA in proliferating myoblasts, we prepared proliferating myoblast-specific, subtracted cDNA probes to screen a myocyte cDNA library. Based on a number of criteria, three cDNAs were selected and characterized. We show that expression of one of these cDNAs transfected into C3HlOT112 fibroblasts, where it is not normally expressed, is sufficient to convert them to stable myoblasts. Myogenesis also occurs, but to a lesser extent, when this cDNA is expressed in a number of other cell lines. The major open reading frame encoded by this cDNA contains a short protein segment similar to a sequence present in the myc protein family.