Oocyte MI completion is prevented by the loss of Cen-RNA. (A) A Cen-ASO was designed against the MinSAT transcripts. (B) The knockdown efficiency of Cen-ASO was tested by qRT-PCR (Cen-ASO: 20, 40, and 100 µM; 5MM-ASO: 40 µM). Analysis of qRT-PCR from three independent experiments (*, P < 0.05; **, P < 0.01; 95% confidence interval, 5-MM: 0.86-1.14; 20 µM: 0.68-0.82; 40 µM; 0.35-0.61; 100 µM: 0.01-0.07). 30 oocytes in each group. (C) Percentage maturation rate following Cen-ASO injection at concentrations indicated, or with water or 5MM-ASO (*, P < 0.05; ****, P < 0.0001; χ2 test). Error bars indicate SD. (D) The timing of first polar body extrusion (PBE) in Cen-ASO–injected oocytes, as indicated, with reversine (100 nM) added at NEBD (****, P < 0.0001; ANOVA with Tukey’s post hoc test). Error bars indicate SD. (E) Timing of meiotic maturation in oocytes injected as labeled. (F) Representative time-lapse images of securin-YFP (gray) expression in oocytes injected with Cen-ASO or 5MM-ASO. Reversine (100 nM) was added at 6 h after NEBD. Scale bar: 20 µm. Times are from NEBD. (G) Securin degradation rates (securin-YFP intensity compared with initial time point, Ft/F0 ) in 5MM-ASO– and Cen-ASO–injected oocytes. Reversine was added as in F. The number of oocytes analyzed in three independent experiments is indicated. Error bars are SD.
