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学科主题: 物理化学
题名: Construction of a high-efficiency multi-site-directed mutagenesis
作者: Tan, Haidong2;  Li, Yueguang3;  Chen, Ling1;  Kasai, Tomonari1;  Seno, Masaharu1
通讯作者: MasaharuSeno
关键词: Site-directed multibase mutagenesis ;  shift primer ;  hexapeptide gene ;  enterokinase gene
刊名: AFRICAN JOURNAL OF BIOTECHNOLOGY
发表日期: 2011-01-17
卷: 10, 期:3, 页:449-452
收录类别: SCI
文章类型: Article
部门归属: 18
项目归属: 1816
产权排名: 1,1
WOS标题词: Science & Technology ;  Life Sciences & Biomedicine
类目[WOS]: Biotechnology & Applied Microbiology
摘要: Construction of a high-efficiency multi-site-directed mutagenesis
研究领域[WOS]: Biotechnology & Applied Microbiology
英文摘要: Although site-directed mutagenesis has been used in many fields, it still has low rate of success and high cost because of low-yield target products. A modified method for multi-site-directed mutagenesis was developed with shifted primer design and cold-start polymerase chain reaction (PCR). The developed method was successfully applied to hexapeptide gene synthesis and recombinant enterokinase gene modification in the plasmids pET41a and pET24b-EK. The efficiency was pronounced at a 1:10 molar ratio of 7-base mutant products to 705-bp fragment products as control. Even in a 10-base substitution mutagenic PCR, a 1:50 molar ratio of mutant products to 705-bp fragment products was reached. Meanwhile, the quality of mutants was proved through the transformation efficiency and sequencing. This method was beneficial to prepare high-quality multibase mutagenesis and also implied that large-scale multibase mutagenesis was feasible, efficient, economical, and productive.
关键词[WOS]: PCR
语种: 英语
WOS记录号: WOS:000286397500022
Citation statistics: 
内容类型: 期刊论文
URI标识: http://cas-ir.dicp.ac.cn/handle/321008/115580
Appears in Collections:中国科学院大连化学物理研究所_期刊论文

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作者单位: 1.Okayama Univ, Dept Med & Bioengn Sci, Grad Sch Nat Sci & Technol, Okayama 7008530, Japan
2.Chinese Acad Sci, Dalian Inst Chem Phys, Dept Biotechnol, Dalian 116023, Peoples R China
3.Tianjin 4th Ctr Hosp, Dept Gen Surg, Tianjin 300140, Peoples R China

Recommended Citation:
Tan, Haidong,Li, Yueguang,Chen, Ling,et al. Construction of a high-efficiency multi-site-directed mutagenesis[J]. AFRICAN JOURNAL OF BIOTECHNOLOGY,2011,10(3):449-452.
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