Bay 1816032

All inhibitors used for the mouse oocytes are listed in Supplemental Table S3. 5-Itu (Cayman Chemicals; 0.5 μM) was used to inhibit haspin kinase with ethanol (Fisher) as the control (solvent for 5-Itu). To disrupt Bub1 H2A phosphorylation, we added BAY-1816032 (8 μM; MedChem Express); we used DMSO (Sigma) as control (solvent for BAY-1816032). We used ZM447439 (Tocris Bioscience; 5 μM) as an Aurora B/C inhibitor and DMSO (Sigma) as a control (solvent for ZM447439). For all mouse meiosis I experiments, BAY-1816032 and ZM447439 were added at 0 h when transferring oocytes to CZB, while 5-Itu was added after 5 h from CZB oocyte transfer. For all mouse meiosis II experiments, all drugs were added at 11 h. Oocytes were fixed 8 h after CZB oocyte transfer for meiosis I experiments and after 15 h for meiosis II experiments. We treated all meiosis I mouse oocytes with acidic Tyrode’s solution (Millipore Sigma) to remove the zone pellucida when imaging for Aurora kinase C, kinetochores, and DNA.

All inhibitors used for the mouse oocytes are listed in Table S3. 5-Iodotubercidin (5-Itu; Cayman Chemicals; 0.5μM) was used to inhibit haspin kinase with ethanol (Fisher) as the control (solvent for 5-Itu). To disrupt Bub1 H2A phosphorylation, we added BAY-1816032 (8μM, MedChem Express); we used DMSO (Sigma) as control (solvent for BAY-1816032). We used ZM447439 (Tocris Bioscience, 5μM) as an Aurora B/C inhibitor and DMSO (Sigma) as a control (solvent for ZM447439). For all mouse meiosis I experiments, BAY-1816032 and ZM447439 were added at 0h when transferring oocytes to CZB, while 5-Itu was added after 5hrs from CZB oocyte transfer. For all mouse meiosis II experiments, all drugs were added at 11hrs. Oocytes were fixed 8hrs after CZB oocyte transfer for meiosis I experiments and after 15hrs for meiosis II experiments. We treated all meiosis I mouse oocytes with acidic Tyrode’s solution (Millipore Sigma) to remove the zone pellucida when imaging for Aurora kinase C, kinetochores, and DNA.