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| 一种高效构建多位点突变重组质粒的方法 | |
| 引用本文： | 孙晓梅,赵彦翔,梁文星,刘俊峰,黄金光. 一种高效构建多位点突变重组质粒的方法[J]. 植物病理学报, 2017, 47(3): 289-295 |
| 作者姓名： | 孙晓梅 赵彦翔 梁文星 刘俊峰 黄金光 |
| 作者单位： | 青岛农业大学动漫与传媒学院, 青岛 266109; 青岛农业大学农学与植物保护学院, 青岛 266109; 中国农业大学植物保护学院, 北京 100193 |
| 基金项目： | 国家自然科学基金项目(31471735);国家科技基础条件平台子项目(2005DKA21207-17);山东省泰山学者建设经费 |
| 摘 要： | 定点突变技术是蛋白质结构与功能研究的必要手段之一, 随着蛋白质结构生物学深入发展, 研究中要同时构建多位点、批量表达载体, 研究者基于重叠延伸PCR和双接头PCR方法, 优化建立一种高效构建多位点突变重组表达质粒的方法。 |
| 关 键 词： | 定点突变 重叠延伸PCR 双接头PCR 表达载体 |
A Highly Efficient-Approach for Introduction of the Multiple Site-Directed Mutations in A Recombinant Plasmid |
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| SUN Xiao-mei,ZHAO Yan-xiang,LIANG Wen-xing,LIU Jun-feng,HUANG Jin-guang. A Highly Efficient-Approach for Introduction of the Multiple Site-Directed Mutations in A Recombinant Plasmid[J]. Acta Phytopathologica Sinica, 2017, 47(3): 289-295 | |
| Authors: | SUN Xiao-mei ZHAO Yan-xiang LIANG Wen-xing LIU Jun-feng HUANG Jin-guang |
| Affiliation: | College of Animation and Communication, Qingdao Agricultural University, Qingdao 266109, China; College of Agronomy and Plant Protection, Qingdao Agricultural University, Qingdao 266109, China; Department of Plant Pathology, China Agricultural University, Beijing 100193, China |
| Abstract: | Site-directed mutagenesis has become an indispensable technique for deciphering protein structure and function. To serve new development for protein structural biology, simultaneous construction for batch of recombinant expression vector with multiple sites mutations is required. Here, we describe a procedure for the unidirectional insertion of site-directed mutagenized DNA into an expression vector via overlap extension and sticky-end PCR. This method takes advantage of two classical PCR-based techniques and has improved the efficiency of the construction of recombinant plasmid with multiple points mutation. |
| Keywords: | site-directed mutagenesis overlap extension PCR sticky-end PCR expression vector |
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