Effective Characterization of DNA Ligation Kinetics by High-Resolution Melting Analysis

doi:10.1002/cbic.201900489

	Effective Characterization of DNA Ligation Kinetics by High-Resolution Melting Analysis
	Zhu, Jianming 1; Wang, Jing 1,3; Cheng, Kai 1; Chen, Hui 1; An, Ran 1; Zhang, Yaping 1; Komiyama, Makoto 1; Liang, Xingguo 1,2
	2019-12-05
发表期刊	CHEMBIOCHEM
ISSN	1439-4227
页码	5
摘要	High-resolution melting (HRM) analysis has been improved and applied for the first time to quantitative analysis of enzymatic reactions. By using the relative ratios of peak intensities of substrates and products, the quantitativity of conventional HRM analysis has been improved to allow detailed kinetic analysis. As an example, the ligation of sticky ends through the action of T4 DNA ligase has been kinetically analyzed, with comprehensive data on substrate specificity and other properties having been obtained. For the first time, the kinetic parameters (k(obs) and apparent K-m) of sticky-end ligation were obtained for both fully matched and mismatched sticky ends. The effect of ATP concentration on sticky-end ligation was also investigated. The improved HRM method should also be applicable to versatile DNA-transforming enzymes, because the only requirement is that the products have T-m values different enough from the substrates.
关键词	enzyme catalysis high-resolution melting kinetics ligases quantitative analysis
DOI	10.1002/cbic.201900489
收录类别	SCI
语种	英语
资助项目	Fundamental Research Funds for the Central Universities[201812009] ; Shandong Provincial Natural Science Foundation of China[ZR2019BC096] ; National Natural Science Foundation of China[31571937] ; National Natural Science Foundation of China[31571937] ; Shandong Provincial Natural Science Foundation of China[ZR2019BC096] ; Fundamental Research Funds for the Central Universities[201812009]
WOS研究方向	Biochemistry & Molecular Biology ; Pharmacology & Pharmacy
WOS类目	Biochemistry & Molecular Biology ; Chemistry, Medicinal
WOS记录号	WOS:000500756500001
出版者	WILEY-V C H VERLAG GMBH
引用统计	被引频次：4[WOS] [WOS记录] [WOS相关记录]
文献类型	期刊论文
版本	出版稿
条目标识符	http://ir.qdio.ac.cn/handle/337002/163823
专题	海洋生态与环境科学重点实验室
通讯作者	Wang, Jing; Liang, Xingguo
作者单位	1.Ocean Univ China, Coll Food Sci & Engn, Qingdao 266003, Peoples R China 2.Qingdao Natl Lab Marine Sci & Technol, Lab Marine Drugs & Bioprod, Qingdao 266003, Peoples R China 3.Chinese Acad Sci, Inst Oceanol, CAS Key Lab Marine Ecol & Environm Sci, Qingdao 266071, Peoples R China
通讯作者单位	海洋生态与环境科学重点实验室
推荐引用方式 GB/T 7714	Zhu, Jianming,Wang, Jing,Cheng, Kai,et al. Effective Characterization of DNA Ligation Kinetics by High-Resolution Melting Analysis[J]. CHEMBIOCHEM,2019:5.
APA	Zhu, Jianming.,Wang, Jing.,Cheng, Kai.,Chen, Hui.,An, Ran.,...&Liang, Xingguo.(2019).Effective Characterization of DNA Ligation Kinetics by High-Resolution Melting Analysis.CHEMBIOCHEM,5.
MLA	Zhu, Jianming,et al."Effective Characterization of DNA Ligation Kinetics by High-Resolution Melting Analysis".CHEMBIOCHEM (2019):5.

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