The objective of this study was to examine the relationship between the timing of the first cleavage and in vitro development of somatic cell nuclear transfer (SCNT) embryos produced by different activation protocols. SCNT embryos were activated with calcium ionophore A23187 and further treated with 6-dimethylaminopurine (DMAP group), cycloheximide (CHX group), or anisomycin (ANI group). The proportion of SCNT embryos that cleaved within 18 hours after activation was significantly higher in the DMAP group (20%) than that in the CHX and ANI groups (3% and 2%, respectively). More than 70% of the cleaved embryos were observed within 24 hours in the DMAP and CHX groups, and within 26 hours in the ANI group. The blastocyst formation rate of SCNT embryos decreased gradually as the time from activation to the first cleavage increased in the DMAP group. The blastocyst formation rate of SCNT embryos cleaved at 22 hours (>20 to ≤22 hours) in the CHX group or within 26 hours in the ANI group was significantly higher than that of SCNT embryos cleaved more than 26 hours in each group. These results indicate that the activation protocol affects the timing of the first cleavage and subsequent in vitro development potential of bovine SCNT embryos and that late-cleaving embryos have a low developmental potential irrespective of the activation protocol.