Polo-like kinase 1 (Plk1) has multiple roles in somatic cell and mammalian oocyte division. In mice, Plk1 distributes to the centromeres from prophase to anaphase and compose spindle apparatus in mitosis stages. Somatic cell nuclear transfer (SCNT) has diverse advantages. However, low cloning efficiency of SCNT procedure causes difficulty to application. The causes of this low efficiency are still unclear. However, they are attributed to the cumulative results of several biological and technical factors. In this study, Plk1, a biological factor, was investigated. B6D2F1 mice (7 weeks old) were superovulated with 10 IU of pregnant mare’s serum gonadotropin and 9 U of human chorionic gonadotropin (HCG) 48 hr later. The oocytes were collected 14 hr after HCG injection and cultured on potassium simplex optimized medium. The BI2536, Plk1-specific inhibitor, was used to understand the influence of Plk1. Also, the embryos were assessed by immunofluorescence. All BI2536-treated embryos failed to the first mitotic division. It showed Plk1 has a critical role in the first mitotic division of the mouse embryo. Moreover, there were significant differences between the control and SCNT embryos in the patterns of Plk1. All SCNT embryos which failed 2-cell development presented incorrect positioning and low expression of Plk1. On the other hand, the control embryos which failed to 2-cell division showed only low expression of Plk1. Taken together, this results demonstrate that Plk1 is critical for successful mitotic division of mouse embryos. Also, correct localization of Plk1 has crucial effect in the development of murine SCNT embryos.