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| Over-expression of Mitofusin 2 Gene(Mfn2) Has Effect on Mouse(Mus musculus) Oocytes In vitro Maturation |
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Abstract Mitofusin 2(Mfn2) participates in apoptosis and cell cycle regulation, which also has an important role in oocyte and embryo development. In order to determine Mfn2 gene effect on mouse(Mus musculus) oocytes during in vitro maturation(IVM) in the present study, firstly, the Mfn2 gene was cloned from mouse ovary by RT-PCR, and an eukaryotic expression vector pMfn2-Venus was constructed. pMfn2-Venus was transfected into 293T cells(virus pakge cell) after mediation by Lipofectamine 2000, identified by fluorescence microscopy observation and Real-time PCR. Then, the cRNA of Mfn2-Venus transcriped in vitro was microinjected into mouse oocytes, and the expression and location was observed under fluorescence microscopy. Finally, the oocytes which were injected Mfn2-Venus cRNA were collected for mature cultivation and the rate of germinal vesicle break down(GVBD) and first polarbody(PB1) were calculated, respectively. The results showed that the recombinant vector pMfn2-Venus was successfully constructed. After transfection or microinjection, the fusion protein could express efficiently and localize accurately both in 293T cells and mouse oocytes. This greatly facilitated the further study of Mfn2, especially its role on oocytes maturation and early embryo development. The rate of GVBD and PB1 were reduced significantly(P＜0.0001, P＜0.05) compared with the that of control group. In conclusion, these data demonstrated that Mfn2 gene had effect on mouse oocytes maturation, it provides a new direction in the research of in vitro maturation of oocytes and a platform for the study of the Mfn2 gene in the process of meiosis.
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Received: 04 August 2014
Published: 13 January 2015
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