|
|||||
|
|
| 改进重叠延伸法引入DNA定点突变的新方法 | |
| 引用本文： | 郭志义,郝小惠,延晋雷,尚利梅,裴鑫,武慧婧,赵偲宇,张璇,杨方.改进重叠延伸法引入DNA定点突变的新方法[J].生物技术,2009,19(6):34-36. |
| 作者姓名： | 郭志义 郝小惠 延晋雷 尚利梅 裴鑫 武慧婧 赵偲宇 张璇 杨方 |
| 作者单位： | 1. 华北煤炭医学院河北省煤矿卫生与安全重点实验室;华北煤炭医学院实验研究中心,河北唐山,063000 2. 华北煤炭医学院实验研究中心,河北唐山,063000 3. 华北煤炭医学院生物学系,河北唐山,063000 |
| 摘 要： | 目的：改进重叠延伸PCR法,实现一种引入DNA定点突变的准确简便方法。方法：通过应用不同的扩增酶和反应体系,以重叠延伸PCR的方法产生引入突变位点的DNA片断,然后再亚克隆到载体中。该文以人cyclin D1启动子的NF-κB位点（-39/-30）为例。结果：通过DNA测序证明定点突变成功引入。一次引入4个突变碱基。突变引入率为100%。 |
| 关 键 词： | 定点突变 重叠延伸PCR 高GC含量 |
A New Method of Site-directed Mutagenesis by Modifying Overlap Extension PCR |
|
| GUO Zhi-yi,HAO Xiao-hui,YAN Jin-lei,SHANG Li-mei,PEI Xin,WU Hui-jing,ZHAO Si-yu,ZHANG Xuan,YANG Fang.A New Method of Site-directed Mutagenesis by Modifying Overlap Extension PCR[J].Biotechnology,2009,19(6):34-36. | |
| Authors: | GUO Zhi-yi HAO Xiao-hui YAN Jin-lei SHANG Li-mei PEI Xin WU Hui-jing ZHAO Si-yu ZHANG Xuan YANG Fang |
| Abstract: | Objective: To develop a new method for generation of site-directed mutation by modifying overlap extension PCR. Method: mutation generation by overlap extension PCR using different DNA polymerase and reaction buffer. The mutation fragment is subcloned into T-vector. In this study, we selected the NF-κB site(-39/-30)of human cyclin D1 promoter for example. Result: The mutation is verified by DNA sequencing. Four point-mutations were intruduced by one-step. The overall rate of obtaining the multiple mutant sites was 100% . Conclusion: An effective and cheap method of DNA mutation by modifying overlap extension PCR had developed. |
| Keywords: | site-directed mutagenesis overlap extension PCR GC rich |
| 本文献已被 维普 万方数据 等数据库收录！ | |