The present experiment was carried out to evaluate the effect of coculturing on myogenic and adipogenic marker gene expressions with the use of C2C12 and 3 T3-L1 preadipocyte cells under the coculture system. C2C12 and 3 T3-L1 cells were cocultured using transwell inserts with a 0.4-mu m porous membrane to separate C2C12 and 3 T3-L1 cells. Each cell type was grown independently on the transwell plates. Following cell differentiation, inserts containing 3 T3-L1 cells were transferred to C2C12 plates, and inserts containing C2C12 cells were transferred to 3 T3-L1 plates. After coculture of the C2C12 and 3 T3-L1 cells for 48 and 72 h, the cells in the lower well were harvested for analysis, and this process was carried out for both cells. Myogenic markers such as myogenin, MyoD, Myf5, PAX3, and PAX7 mRNA expressions were analyzed in the cocultured C2C12 cells. Adipogenic markers such as fatty acid-binding protein 4 (FABP4), peroxisome proliferator-activating receptor (PPAR gamma), CCAAT/enhancer-binding protein (CEBPA), adiponectin, lipoprotein lipase, and fatty acid synthase mRNA expressions were analyzed in the cocultured 3 T3-L1 cells. Myogenic and adipogenic marker gene mRNA expressions were significantly altered in the cocultured C2C12 and 3 T3-L1 cells when compared with the monocultured C2C12 and 3 T3-L1 cells.