目录
摘 要 III
关键词 III
Abstract V
引言
引言 1
1 材料与方法 3
1.1 试验材料 3
1.1.1 抗体和试剂 3
1.1.2 主要仪器和设备 3
1.1.3 试剂的配制 3
1.2 试验方法 4
1.2.1 小鼠GV期卵母细胞的采集和体外培养 4
1.2.2 小鼠成熟卵母细胞的采集 4
1.2.3 小鼠卵母细胞的体外受精 5
1.2.4 猪卵母细胞的收集和体外培养 5
1.2.5 猪卵母细胞的体外受精 6
1.2.6 免疫荧光染色&共聚焦显微成像 6
2 结果与分析 ７
2.1 在猪卵母细胞中Juno是Izumo1的受体 ７
2.2 小鼠卵母细胞发育过程中Juno的定位 ７
2.3 小鼠受精过程以及早期胚胎发育过程中Juno的定位 ８
2.4 猪卵母细胞发育过程中Juno的定位 ９
2.5 猪卵母细胞受精过程以及早期胚胎发育过程中的定位 １０
3 讨论 １２
致谢 １３
参考文献 １３
Juno在哺乳动物卵子受精后的动态变化研究
摘 要
多精受精现象会导致动物胚胎发育异常和家畜繁殖力低下，是制约畜牧业发展的重要原因之一。与其他哺乳动物相比，猪的多精受精发生率很高，特别在体外受精过程中，但其具体的发生机理目前仍不清楚。为了探究精卵融合蛋白Juno在哺乳动物卵母细胞受精后的动态变化是否能为解析多精受精现象提供科学依据，本研究利用Juno、Izumo1重组蛋白和抗体，通过免疫荧光染色，观察Juno蛋白在猪和小鼠卵母细胞减数分裂成熟过程、受精过程和早期胚胎发育时期的动态变化规律，分析比较二者的异同，明确其在精卵融合和预防多精受精中的作用。结果表明，在小鼠上，膜蛋白Juno在受精后短时间内便已从卵母细胞质膜上消失，而猪卵子膜蛋白Juno在减数分裂、受精过程中均正常表达，受精后短时间内并没有快速 *51今日免费论文网|www.51jrft.com +Q: ￥351916072$ 
消失，甚至到四细胞时期仍有较高水平的表达。由此，我们推断猪多精受精现象发生率较高与受精后Juno未从卵子质膜上快速消失有关，从而未能有效使卵质膜发生质膜屏蔽阻止多精受精。
THE STUDY ON THE DYNAMIC CHANGES OF JUNO IN MAMMALIAN EGGS AFTER FERTILIZATION
ABSTRACT
Polyspermy is one of the important causes leading to the abnormal development of animal embryos and low fertility of livestock, thereby restricting the development of animal husbandry. Compared with other mammals, the incidence of polyspermy in pigs is dramatically high, especially during in vitro fertilization. However, its detailed mechanism is still unclear. To ask whether the dynamic changes of spermegg fusion protein Juno after fertilization in mammals could provide a scientific evidence for interpreting the phenomenon of polyspermy, our study used Juno and Izumo1 recombinant proteins and antibodies to observe the dynamic patterns of Juno during oocyte maturation, fertilization process and early embryonic development in porcine and mouse oocytes by immunostaining. We compared the similarities and differences of Junos dynamics between two species, and clarify their roles in spermegg fusion and prevention of polyspermy. The results showed that the fusion protein Juno disappeared from oocyte membrane within a short time after fertilization in mice. By contrast, Juno persisted on the membrane after fertilization in porcine oocytes. Even in the porcine fourcell embryos, there was still a high level of Juno. From this observation, we inferred that the high incidence of polyspermy in pigs is related to the delayed disappearance of Juno from the egg plasma membrane after fertilization, thus failing to activate the membrane block to polyspermy.

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