The mouse mdr1a and mdr1b genes are expressed in skeletal muscle, though their precise role in muscle is unknown. Dystrophic muscle is characterized by repeated cycles of degeneration and regeneration. To explore the role of the mdr1 genes during muscle regeneration, we have created a triple knockout mouse lacking the mdr1a, mdr1b, and the dystrophin genes. The resulting ReX mice developed normally and were fertile. However, as adults, ReX had a higher proportion of degenerating muscle fibers and greater long-term loss of muscle mass than mdx. ReX muscles were also characterized by a reduced proportion of muscle side population (mSP) cells, of myogenic cells, and a reduced capacity for muscle regeneration. We found too that mSP cells derived from dystrophic muscle are more myogenic than those from normal muscle. Thus, in dystrophic muscle, the mdr1 gene plays an important role in the preservation of the mSP and of the myogenic regenerative potential. Moreover, our results suggest a hitherto unappreciated role of mdr1 in precursor cells of regenerating tissue; they therefore provide an important clue to the physiological significance of mdr1 expression in stem cells.