Microinjection instrument

All injections were carried out according to the procedures described previously36 (link). The small interfering RNA (siRNA) of Mfn2 (sequence: UCCUCAAGGUUUAUAAGAATT) (GenePharma, Shanghai, China), or the siRNA control, was microinjected (25 μM) into the cytoplasm of fully grown GV oocytes with an Eppendorf microinjection instrument (Hamburg, Germany) and completed within 30 min. Oocytes were kept in M2 medium supplemented with 2.5 μM milrinone (Sigma-Aldrich, St. Louis, MO, USA) to prevent GV breakdown and to allow Mfn2-siRNA to complete its role during this period. After 24 h, the oocytes were cleaned thoroughly to resume meiosis. Each experiment was repeated three to five times.

Small interfering RNAs (siRNA) for Micu1 (sequence: AGCCUUAUCCUGAGGACAATTU UGUCCUCAGGAUAAGGCUTT), Micu2 (sequence: CCUCUUCUCAGUCAUGUUUTTAAACAUGACUGAGAAGAGGTT), NCLX (sequence: CCUUCUUGCCACGUCUAATTUUAGACGUGGCAAAGAAGGTT), MCU (sequence: CCAAAGAGACCUAATTUUAGGAGGUCUCUCUUUGGTT) (GenePharma, Shanghai, China), or siRNA control were microinjected (5 μM) into the cytoplasm of fully grown GV oocytes with an Eppendorf microinjection instrument (Hamburg, Germany) and completed within 30 min. Oocytes were arrested in M16 supplemented (Sigma-Aldrich, St. Louis, MO, USA) with 2.5 μM milrinone for 20 h to block mRNA translation. After 20 h, the oocytes were cleaned thoroughly to resume meiosis.

Small interference RNAs (siRNA) for MCU (CCAAAGAGACCUAATTUUAG GAGGUCUCUCUUUGGTT) and
Mad2 (GGACUCACCUUGCUUACAATTUUGUAAGCAAGGUGAGUCCTT) (Gene Pharma) or siRNA‐negative controls were microinjected (5 μM) into fully grown immature oocytes with an Eppendorf microinjection instrument (Hamburg) and allowed to incubate for 30 min. Oocytes were arrested in the GV stage in MI6 medium (Sigma‐Aldrich) supplemented with 2.5 μM milrinone for 20–24 h. The oocytes were then thoroughly washed with DPBS to resume meiosis.