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After microscopic examination, oocytes at metaphase II stages (MII, with the polar body extrusion) were collected and cultured in 500 l of fertilization droplets as SOF medium supplemented with 5% fetal bovine serum, 2 mM sodium pyruvate, 10 g/ml heparin sodium and 1 mM caffeine in a CO 2 incubator (38.5C, 90%N 2 , 5%CO 2 and 5%O 2 ) ( For in vitro fertilization (IVF), the frozen-thawed semen of each group was placed in the 15 ml tube containing the Percoll density gradient