An improved PCR-based method for site directed mutagenesis using megaprimers
An improved PCR-based method for site directed mutagenesis using megaprimers
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DOI：
10.1006/mcpr.1998.0187
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发表时间：
1998-12-01
影响因子：
3.3
通讯作者：
Kristiansen, K
中科院分区：
文献类型：
--
作者：
Brons-Poulsen, J;Petersen, NE;Kristiansen, K
介绍了一种基于两步聚合酶链反应（PCR）巨引物方法的位点定向诱变改进方案。与以前发表的方案相比，本文中描述的方案确保了至少85%的成功率，基本上没有引入不必要的二次突变。该方案的基本特点包括优化模板-引物的数量和比例，允许使用减少的PCR循环次数和使用校对耐热DNA聚合酶。(C) 1998学术出版社。
An improved protocol for site-directed mutagenesis based on the two-step polymerase chain reaction (PCR) megaprimer method is described. Compared to previously published protocols, the protocol described in this article ensures consistently a success rate of at least 85% with essentially no introduction of unwanted secondary mutations. The essential features of this protocol include an optimization of the template-primer amounts and ratio that allows the use of a reduced number of PCR cycles and the use of proof-reading thermostable DNA polymerases. (C) 1998 Academic Press.