安徽农业科学2012，Vol.40Issue(10)：5779-5781,3.
利用重叠延伸PCR技术进行定点突变研究
Site-directed Mutagenesis Based on Overlap Extension PCR
摘要
Abstract
[ Objective ] To establish an efficient, convenient and economical method for site-directed mutagenesis. [ Method ] The target mutation was introduced into primers designed by DNAMAN5.0 software. Through overlap extension PCR for twice obtained the mutation gene which of the full length of the recombinant Human Tissue type plasminogen activator ( Reteplase ). The mutation gene cloned it into Peasy-blunt simple cloning vector for sequencing. [ Result ] The sequencing results showed that three site mutations were fully consistent with the expected results (10th site had been added a base-pair of A, C had been changed into G at 137th site, G had been changed into A at 686th site). Three site mutations were introduced by using overlap extension PCR on one-step. The overall rate of obtaining the mutant sites was 100%. Site-directed mutagenesis will clone the recombinant Human Tissue type plasminogen activator and laid the basis for the functional study. [Conclusion] Site-directed mutagenesis was successfully implemented based on the overlap extension PCR which is an efficient, convenient and economical DNA-directed mutagenesis method.关键词
重叠延伸PCR技术/定点突变/rPA基因Key words
Overlap extension PCR/ Site-directed mutagenesis/ Human Tissue Plasminogen Activator (Reteplase)分类
农业科技引用本文复制引用
雒丽娜,王盛,王玉炯..利用重叠延伸PCR技术进行定点突变研究[J].安徽农业科学,2012,40(10):5779-5781,3.基金项目
国家自然科学基金资助项目(31160032). （31160032）
