An efficient approach for site-directed mutagenesis using central overlapping primers
An efficient approach for site-directed mutagenesis using central overlapping primers
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使用中心重叠引物进行定点诱变的有效方法
DOI：
10.1016/j.ab.2011.07.008
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发表时间：
2011-11-15
影响因子：
2.9
通讯作者：
Liu, Runzhong
中科院分区：
文献类型：
--
作者：
Wang, Huiping;Zhou, Nan;Liu, Runzhong
QuikChange是蛋白质和核酸结构和功能研究中常用的定点突变方法。然而，标准方案在产生所需突变方面通常效率低下。在这里，我们提出了一种新的引物设计策略，中央重叠引物（COP），它采用了一对不同长度的二分引物，与长引物的中间区域互补的短引物。COP方法在产生约90%的突变率方面是有效和稳健的，而无需超感受态大肠杆菌细胞或费力地筛选阳性克隆。(C)2011 Elsevier Inc. All rights reserved.
QuikChange is a popular method for site-directed mutagenesis in structural and functional studies of proteins and nucleic acids. However, the standard protocol is often inefficient in producing the desired mutations. Here we present a novel strategy for primer design, central overlapping primers (COP), which employs a pair of bipartite primers of different lengths, with the short primer complementary to the middle region of the long primer. The COP method is efficient and robust in generating approximately 90% mutation rate without supercompetent Escherichia coli cells or laborious screening for positive clones. (C) 2011 Elsevier Inc. All rights reserved.