FIGURE 2.

Western blot analysis of Cdc25B protein expression and phosphorylation status of Cdc2-Tyr¹⁵, cleavage rate, and MPF activity after mRNAs encoding Cdc25B-WT, Cdc25B-S149A, Cdc25B-S321A, Cdc25B-S149A/S321A, Cdc25B-S149D, Cdc25B-S321D, or Cdc25B-S149D/S321D were microinjected into fertilized mouse eggs in the absence of dbcAMP and H-89 in M16 medium. A, Western blot analysis of Cdc25B protein expression at 4 h after microinjection of Cdc25B mRNAs. B, the cleavage rate in cultured mouse embryos of various Cdc25B mRNAs injection at 31 h after hCG injection. The cleavage rates were calculated with data from three independent experiments. C, Western blot analysis of the phosphorylation status of Cdc2-Tyr¹⁵ in the various Cdc25B mRNA injection groups. The eggs were collected at 26.5, 27, 27.5, 28, 28.5, and 29 h after hCG injection. D, MPF activity in the control groups, including no injection and TE buffer groups. E, MPF activity in eggs injected with Cdc25B-WT-mRNA. F, MPF activity in eggs injected with Cdc25B-S149A-mRNA. G, MPF activity in eggs injected with Cdc25B-S321A-mRNA. H, MPF activity in eggs injected with Cdc25B-S149A/S321A-mRNA. I, MPF activity in eggs injected with Cdc25B-S149D-mRNA. J, MPF activity in eggs injected with Cdc25B-S321D-mRNA. K, MPF activity in eggs injected with Cdc25B-S149D/S321D-mRNA. For each point, five eggs were collected, and MPF activity was examined by scintillation counting and autoradiography. Each value was expressed as the mean ± S.D. from three independent experiments. A total of ∼200 eggs were loaded onto each lane in Western blot.