人工锌指核酸酶介导的基因组定点修饰技术-CALIS机构知识库.PDFVIP

HEREDITAS (Beijing) 2011 7 , 33(7): 665―683 ISSN 0253-9772 综述 DOI: 10.3724/SP.J.1005.2011.00665 人工锌指核酸酶介导的基因组定点修饰技术 肖安, 胡莹莹, 王唯晔, 杨志芃, 王展翔, 黄鹏, 佟向军, 张博, 林硕 , 100871 锌指核酸酶(ZFN) 由锌指蛋白(ZFP)结构域和Fok I 核酸内切酶的切割结构域人工融合而成, 是近年来发 展起来的一种可用于基因组定点改造的分子工具。ZFN 可识别并结合特定的DNA 序列, 并通过切割这一序列的特 定位点造成DNA 的双链断裂(DSB)。在此基础上, 人们可以对基因组的特定位点进行各种遗传操作, 包括基因打靶、 基因定点插入、基因修复等, 从而能够方便快捷地对基因组实现靶向遗传修饰。这种新的基因组定点修饰方法的突 出优势是适用性好, 对物种没有选择性, 并且可以在细胞和个体水平进行遗传操作。文章综述了ZFN 技术的研究进 展及应用前景, 重点介绍ZFN 的结构与作用机制、现有的靶点评估及锌指蛋白库的构建与筛选方法、基因组定点修 饰的策略, 以及目前利用这一技术已成功实现突变的物种及内源基因, 为开展这一领域的研究工作提供参考。 锌指核酸酶; 锌指蛋白; 基因组定点修饰; 基因打靶; 分子工具 Progress in zinc finger nuclease engineering for targeted genome modification XIAO An, HU Ying-Ying, WANG Wei-Ye, YANG Zhi-Peng, WANG Zhan-Xiang, HUANG Peng, TONG Xiang-Jun, ZHANG Bo, LIN Shuo Key Laboratory of Cell Proliferation and Differentiation of Ministry of Education, College of Life Sciences, Peking University, Beijing 100871, China Abstract: Zinc finger nuclease (ZFN) is an artificially engineered hybrid protein that contains a zinc finger protein (ZFP) domain and a Fok I endonuclease cleavage domain. It has recently emerged as a powerful molecular tool for targeted ge- nome modifications. ZFNs recognize and bind to specific DNA sequences to generate a double-strand break (DSB) by its nuclease activity. Based on this finding, various genetic methods, including gene targeting (gene disruption), gene addition, gene correction etc., are being designed to manipulate the genomes of different species at specific loci. One particular ad- vantage of this new technique is its broad