Objective To observe the effects of ubiquitin C-terminal hydrolase L1 (UCHL1) mutant protein and wild-type protein overdose on mouse oocyte maturation, and analyze its role and interaction mode in mouse oocyte maturation in vitro. Methods The fusion proteins of UCHL1 mutations (C90S and I93M) and wild-type UCHL1 proteins with glutathione S-transferase (GST) were prepared by recombinant protein expression technology. The fusion proteins, GST and phosphate buffered solution (PBS) were injected into immature oocytes by microinjection technology, or wild-type proteins were added to in vitro culture medium to analyze the effects of wild-type UCHL1 overdose, UCHL1 (I93M) mutant protein addition or addition of UCHL1 (C90S) mutant protein with loss of ubiquitin hydrolase activity on oocyte germinal vesicle breakdown (GVBD). The germinal vesicle oocytes of UCHL1 (I93M) mutant mice and wild-type mice were isolated and cultured in vitro for 3 h, and then the GVBD rates were compared. Results There was no significant difference in the GVBD rates of oocytes between microinjection groups with UCHL1- or its mutants-GST fusion protein, GST injection group or PBS injection group (all P<0.05). There was also no significant difference in the GVBD rates of oocytes in the group added with UCHL1-GST fusion protein in the culture medium compared with the control group without injection or addition (P>0.05). A small number of germinal vesicle oocytes in the UCHL1 (C90S)-GST fusion protein injected group progressed to MⅡ stage oocytes in vitro, and the polar body was larger than that of the control group. There was no significant difference in GVBD rates of germinal vesicle oocytes in vitro between UCHL1 (I93M) mutant mice and wild-type mice (P>0.05). Conclusion The addition of exogenous UCHL1, UCHL1 (I93M) mutant with toxic effects, or UCHL1 (C90S) mutant with loss of hydrolase activity in mouse oocytes do not affect the GVBD process of oocyte maturation. The GVBD rate of mouse oocytes with UCHL1 (I93M) mutation is normal. However, the mutation of UCHL1 (C90S) may affect the formation of polar bodies.