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哺乳动物雌性生殖细胞及早期胚胎基因敲除或敲减的方法

张美玲¹, 郝鑫¹, 周成杰¹, 王璐¹, 宋春儒¹, 韩哲¹, 张晓洁¹, 张瑞丰¹, 梁成光^1,*

¹内蒙古大学省部共建草原家畜生殖调控与繁育国家重点实验室，实验动物研究中心，生命科学学院，呼和浩特 010070

摘要

哺乳动物雌性生殖细胞为胚胎的形成和发育提供了大量母源性遗传物质。受精一旦实现，所形成的早期胚胎通过细 胞分裂与分化逐渐形成具有复杂结构的全新个体。针对雌性生殖细胞(主要是卵母细胞)的相关研究既能解析其独有的生理学 特性，又能为其他类型细胞的相关研究提供借鉴作用。然而，哺乳动物卵母细胞数量稀少，且不能在体外进行扩增培养， 导致针对卵母细胞的相关研究难度极大。哺乳动物早期胚胎的研究也同样面临材料短缺的问题。在以卵母细胞和早期胚胎 为材料的研究中，针对特定基因功能的失活或干扰是最常用的方法之一。本文将结合他人和本研究组在哺乳动物雌性生殖 细胞及早期胚胎中进行的基因功能敲除或敲减实验，对条件性基因敲除、RNA干扰、Morpholino、Trim-Away和抗体介导的 蛋白功能干扰这几类技术的主要方法原理、案例、优缺点和使用注意事项进行总结分析，并提出未来相关技术可能的改进 方向，以期为从事卵母细胞及早期胚胎发育研究的科技工作者提供参考。

关键词： 哺乳动物; 卵母细胞; 早期胚胎; 敲除; 敲减

分类号：Q954.43+2;Q786

Methods used for gene manipulation in mammal female germ cells and early embryos

ZHANG Mei-Ling¹, HAO Xin¹, ZHOU Cheng-Jie¹, WANG Lu¹, SONG Chun-Ru¹, HAN Zhe¹, ZHANG Xiao-Jie¹, ZHANG Rui-Feng¹, LIANG Cheng-Guang^1,*

¹State Key Laboratory of Reproductive Regulation & Breeding of Grassland Livestock, The Research Center for Laboratory Animal Science, School of Life Sciences, Inner Mongolia University, Hohhot 010070, China

Abstract

For sexual reproduction, oocytes are mammalian female germ cells that provide the majority of maternal genetic material for early stage embryo production and development. Early stage embryos begin the process of multicellular organism formation through cell differentiation. Studies on mammalian female germ cells (oocytes) not only reveal its unique physiological characteristics, but also help understand the mechanism involved in cell differentiation of other cell types. However, because it is difficult to culture in vitro, our understanding of the function of oocytes and early stage embryos remains very limited. Gene editing or manipulation is one of the most commonly used method, which is also useful in the field of gametes study. In this review, we summarized the principles, advantages and disadvantages of techniques, which include conditional knockout, RNA interference, Morpholino, Trim-Away and antibody-mediated inhibition of protein function, currently used for gene manipulation in oocytes and early stage embryos. We also discuss the issues the investigators need to consider. Finally, we highlight the future directions for gene manipulation or editing in female germ cells and early stage embryos.

Key words: mammal; oocytes; early stage embryos; knockout; knockdown

收稿日期：2019-06-30　　录用日期：2019-09-12

通讯作者：梁成光　　E-mail: liangcg@imu.edu.cn

DOI: 10.13294/j.aps.2019.0081

引用本文：

张美玲, 郝鑫, 周成杰, 王璐, 宋春儒, 韩哲, 张晓洁, 张瑞丰, 梁成光. 哺乳动物雌性生殖细胞及早期胚胎基因敲除或敲减的方法[J]. 生理学报 2020; 72 (1): 31-47.

ZHANG Mei-Ling, HAO Xin, ZHOU Cheng-Jie, WANG Lu, SONG Chun-Ru, HAN Zhe, ZHANG Xiao-Jie, ZHANG Rui-Feng, LIANG Cheng-Guang. Methods used for gene manipulation in mammal female germ cells and early embryos. Acta Physiol Sin 2020; 72 (1): 31-47 (in Chinese with English abstract).