To investigate the roles played by MyoD in the terminal differentiation of satellite cell‐derived myoblasts, the effect of antisense inhibition of MyoD expression was examined in bovine adult myoblast culture, in which inhibition treatment was limited to the terminal differentiation phase. MyoD antisense oligonucleotide DNA (AS‐mD) suppressed the formation of multinucleated myotubes in the cell culture. Myotube formation was suppressed even when AS‐mD treatment was limited to the period preceding the onset of myotube formation. Reverse transcriptase–polymerase chain reaction (RT–PCR) analysis revealed that treatment with AS‐mD suppressed the expression of myosin heavy chain embryonic isoform and troponin T isoforms at 4 days after the induction of differentiation. AS‐mD also suppressed the expression of MRF4, but did not alter the expression of either Myf5 or myogenin, in contrast to previous results using mouse cells possessing MyoD(–/–) genetic background. These findings suggest that MyoD controls myogenesis but not Myf5 or myogenin mRNA expression during the terminal differentiation phase. Furthermore, among the α4, α5, α6, and α7 integrins, α4, α5, and α7 integrin expression was suppressed by AS‐mD treatment, in parallel with the suppression of myotube formation, which suggests that MyoD controls myotube formation by regulating the expression of α4, α5, and α7 integrins.