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. 2019 Feb 15;218(5):1553–1563. doi: 10.1083/jcb.201808088

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© 2019 Li et al.

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Figure 3. — Ccnb3 mutation with inefficient APC/C activity and functional SAC. (A–D) Time-lapse fluorescence measurement of Ccnb1-EGFP (A and B) and securin-EGFP (C and D) expressed of indicated mRNA injection at GV stage, respectively. The integrated intensities of EGFP were measured, background corrected, and normalized to the initial-intensity value obtained per oocyte. GV-stage oocytes were injected with 100 ng/µl Ccnb1-EGFP or securin-EGFP mRNA, incubated in M2 medium containing dbcAMP for 2 h; then, oocytes were washed with dbcAMP-free M2 medium to resume meiosis. At least 10 and 20 oocytes for Ccnb1-EGFP and securin-EGFP were tested, respectively. Measurements were aligned to 4 h after GVBD as the starting time. The asterisk indicates the PB1. Statistical analyses for differential securin-GFP fluorescence intensity changes. Error bars represent mean ± SD; ***, P < 0.001 (Student’s t test). (E) Chromosome spreads were prepared 4 h (pro-MetI), 6 h (MetI), and 8 h (AnaI) after GVBD and then stained with SECURIN (red) and Hoechst 33342 (blue). (F) Chromosome spreads were prepared 4 h (pro-MetI) and 6 h (MetI) after GVBD, then stained with BUB3 (green) and Hoechst 33342 (blue). Bars: 20 µm (A and C); 5 µm (E and F).