Full-Text Search:
Home|Journal Papers|About CNKI|User Service|FAQ|Contact Us|中文
《China Biotechnology》 2015-05
Add to Favorite Get Latest Update

A Simple and Rapid Single Primer PCR Method for Site-directed Mutagenesis

GAO Rui-ping;CHENG Long-bin;LI Zhen-qiu;College of Life Sciences,Hebei University;  
To solve the problem that additional primers could be inserted at the mutation site when use Quick Change site-directed mutagenesis methods to mutate some specific sites,Quick Change method was modifed as followed:A pair of complementary primers containing mutations site were synthesized,and single primer PCR amplification was done respectively. The two PCR products from single amplification were mixed and denatured and renatured followed by addition of restriction endonuclease Dpn I. The product was transformed into competent E. coli DH5α,and positive clones were screened for DNA sequencing. Several sites in amorpha-4,11-diene synthase( ADS) gene that could not be successful mutated using tranditional method had been successfully mutated with this method without any additional primers. So this novel method was practicable and could solve the problem of extra primers insertion in Quick Change site-directed mutagenesis. This also indirectly proved that additional primers insertion was caused by putting two primers in one reaction system.
【Fund】: 国家自然科学基金资助项目(30900111)
【CateGory Index】: Q503
Download(CAJ format) Download(PDF format)
CAJViewer7.0 supports all the CNKI file formats; AdobeReader only supports the PDF format.
【Citations】
Chinese Journal Full-text Database 2 Hits
1 LIU Xin-Yi, ZHANG Hui-Zhan, YUAN Qin-Sheng(Department of Biochemistry Engineering, East China University of Science and Technology,Shanghai 200237,China);A Novel Method for Site-directed Mutagenesis and Its Application in DdsA Mutant Research[J];Chinese Journal of Biochemistry and Molecular Biology;2007-05
2 ZHANG Min XIN Shao-jie DUAN Xue-zhang HOU Jun HU Yan Mao Pan-yong Institute of Virology,302th Hospital of People's Liberation Army,Beijing 100039,China;Establishment of a new"twice single primer PCR method"and application for site-directed mutations in HBV pre-core region[J];Chinese Journal of Laboratory Medicine;2007-04
【Co-citations】
Chinese Journal Full-text Database 8 Hits
1 Fanrong MENG;Chen CHEN;Yongwen LI;Haisu WAN;Qinghua ZHOU;Tianjin Key Laboratory of Lung Cancer Metastasis and Tumor Microenviroment, Tianjin Lung Cancer Institute, Tianjin Medical University General Hospital;;A Highly Efficient In Vitro Site-directed Mutagenesis Protocol for Introducing Multiple-site Mutations into Target Genes[J];Chinese Journal of Lung Cancer;2014-06
2 Fanrong MENG;Chen CHEN;Haisu WAN;Qinghua ZHOU;Tianjin Key Laboratory of Lung Cancer Metastasis and Tumor Microenviroment, Tianjin Lung Cancer Institute,Tianjin Medical University General Hospital;;A Method for Introducing Mutations into Large Vectors[J];Chinese Journal of Lung Cancer;2014-07
3 WU Li-shuang;TIAN Jian;ZHENG Jia;GUO Ning;WANG Yu-guang;ZHOU Hong-bo;School of Minerals Processing and Bioengineering,Central South University;;The constitutive expression of gene encoding pullulanase in Pichia pastoris and site-directed mutagenesis[J];Journal of Biology;2013-05
4 ZHAO Juan;WANG Xia;LI Bing-Zhi;CHENG Jing-Sheng;YUAN Ying-Jin;Key Laboratory of System Biological Engineering of the Ministry of Education,School of Chemical Engineering,Tianjin University;;DNA assembly in synthetic biology[J];Chinese Bulletin of Life Sciences;2013-10
5 GUO Zi-Fen;XUAN Gui-Ping;CHEN Fang-Fang;ZHANG Shi-Yi;Institute of Pharmacy and Pharmacology, University of South China;;Construction of Three Point Mutations Vector by Modifying Overlap Extension PCR[J];Letters in Biotechnology;2014-02
6 YANG Hai;WANG Fang-yu;LI Qing-wang;HU Jian-hong;Dong Xiao-xiao;SHAN Yong-hong;Department of Life Science,Hengyang Normal University;College of Animal Science and Technology,Northwest A&F University;Department of Product Research and Development,Nanyue Biopharmaceutical Co.,LTD;;Construction and eukaryotic expression of human antithrombin Cys248 site mutant[J];Journal of Northwest A & F University(Natural Science Edition);2013-10
7 ZHENG Bin;YIN Zhi-kui;YAO Zhi-jun;ZHANG Hai-zhu;REN Hong-bin;ZHAN Xi-mei;Department of Parasitology,Xinxiang Medical University;Department of Pharmacology,Xinxiang Medical University;Department of Parasitology,Zhongshan School of Medicine,Sun Yat-sen University;;Identification of interaction site of MIC2 with aldolase in Toxoplasma gondii[J];Chinese Journal of Zoonoses;2014-07
8 ZHENG Bin;YIN Zhi-kui;ZHAN Xi-mei;Department of Parasitology,Xinxiang Medical University;College of Pharmacy,Xinxiang Medical University;Department of Parasitology,Zhongshan School of Medicine,Sun Yat-sen University;;Protein Interaction Site of Toxoplasma gondii Microneme Protein 6 and Aldolase Determined by Site-directed Mutagenesis[J];Chinese Journal of Parasitology and Parasitic Diseases;2014-03
【Secondary Citations】
Chinese Journal Full-text Database 2 Hits
1 CAO Yang, LI Dongtian, YIN Bingnan, et al. Department of Microbiology, Tianjin Medical University, Tianjin. 300070;Establishment and application of overlap-extension PCR[J];Hebei Medical Journal;2005-11
2 ZENG Jin Yang, HUANG Jia Qiang, LI Zhuo Ya, GONG Fei Li (Department of Immunology, Tongji Medical University, Wuhan 430030);A rapid and simple method of mutagenic deletion[J];JOURNAL OF CELLULAR AND MOLECULAR IMMUNOLOGY;1998-04
©2006 Tsinghua Tongfang Knowledge Network Technology Co., Ltd.(Beijing)(TTKN) All rights reserved