Vertebrate skeletal myofibers are heterogeneous with respect to their metabolic, contractile and morphological properties. To better understand skeletal myofiber diversity and plasticity at the transcriptional level, we carried out a whole-genome gene expression analysis of skeletal muscles composed primarily of slow/oxidative fibers (soleus) and fast fibers (extensor digitorum longus, EDL). We also followed gene expression changes in plantaris muscles from mice undergoing voluntary wheel running, a protocol that triggers transformation of glycolytic fibers into oxidative ones. Microarray analysis identified 70 genes differentially expressed by 3-fold or greater in soleus vs. EDL muscles and 15 genes up-regulated in exercised vs. sedentary plantaris muscles. A subset of these results were verified by northern blot and/or real-time RT-PCR analyses. Our results expand knowledge of the differences among various types of skeletal myofibers and their adaptation to exercise at the transcriptional level.