To reveal the molecular mechanism underlying the membrane protrusions derived from the condensation of homologous chromatin in oocytes induced by demecolcine, the effect of the demecolcine concentration, treatment duration, and synergetic effects with sucrose solution on the rate of membrane protrusions of bovine oocytes were investigated.Using real-time fluorescent quantitative PCR, western blot analysis, and immunofluorescence assays, the expression of the maternal c-mos gene, the activity of mitogen activated protein kinase (MAPK), and the change in the localization of spindles and nuclei during the demecolcine treatment were analyzed in bovine oocytes.Treatment of bovine oocytes with both demecolcine (0.6 μg/mL) and sucrose (0.05 M) for 1 h led to the highest rate of membrane protrusions, and synergistic effects were also observed.Real-time fluorescent quantitative PCR analysis revealed that the demecolcine treatment up-regulated the expression of the maternal c-mos gene.Western blot analysis indicated that the demecolcine and sucrose co-treatment enhanced the activity of MAPK in bovine oocytes.Immunofluorescence analysis indicated that the spindles and nuclei were localized at the place of the membrane protrusions.The mechanism by which demecolcine caused the membrane protrusions may be through the regulation of the expression of c-mos, which then affected the activity of MAPK and eventually caused the change in the morphology and assembly of spindles during meiosis in oocytes.The result provides a theoretical basis for the molecular mechanism underlying chemically assisted enucleation.