高等学校化学学报

高等学校化学学报 ›› 2003, Vol. 24 ›› Issue (10): 1761.

• 论文 • 上一篇    下一篇

基于分子信标实时监测大肠杆菌DNA连接酶催化的DNA连接过程

刘凌凤, 唐志文, 王柯敏, 谭蔚泓, 李军, 郭秋平, 孟祥贤, 黄杉生, 李杜   

  1. 湖南大学化学生物传感与计量学国家重点实验室, 生物技术研究院, 化学化工学院, 长沙 410082
  • 收稿日期:2003-04-15 出版日期:2003-10-24 发布日期:2003-10-24
  • 通讯作者: 王柯敏(1957年出生),男,博士,教授,博士生导师,从事纳米尺度和单分子水平上的生物分析化学研究.E-mail:kmwang@hnu.edu.cn E-mail:kmwang@hnu.edu.cn
  • 基金资助:

    国家重点基础研究发展计划(批准号:2002CB13100);科学技术部基础研究重大项目前期研究专项基金(批准号:2001-51);国家自然科学基金重点项目(批准号:20135010);教育部重大项目基金(批准号:2000-156);海外青年学者合作研究基金(

Real-time Visualization of Ligation Reaction Catalyzed by E.coli DNA Ligase Based on Molecular Beacon

LIU Ling-Feng, TANG Zhi-Wen, WANG Ke-Min, TAN Wei-Hong, LI Jun, GUO Qiu-Ping, MENG Xiang-Xian, HUANG Shan-Sheng, LI Du   

  1. State Key Laboratory of Chemo/Biosensing and Chemometrics, Institute of Biological Technology, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, China
  • Received:2003-04-15 Online:2003-10-24 Published:2003-10-24

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被引次数

19

摘要： 报道了一种对DNA连接过程进行实时监测的方法,利用分子信标核酸探针作为DNA连接反应的模板和检测探针,实时监测了 E.coli DNA连接酶催化的DNA连接反应,克服了传统的凝胶电泳技术操作复杂、周期长及无法实时监测DNA连接过程的缺点,为核酸连接过程的实时监测和连接酶催化机理的研究提供了更为丰富的信息.在此基础上,发展了一种快速、准确测定 E.coli DNA连接酶的方法,线性响应范围为4.0×10-6～2.0×10-4U/μL,检测下限为4.0×10-6U/μL.

关键词: E.coli DNA连接酶, 核酸连接, 分子信标, 实时监测

Abstract: Nucleic acids ligation is a crucial step in replication and repair of DNA and in genetic recombination.Researches on DNA ligation have been generally based on gel electrophoresis and autoradiography in the past, which include complex work and are time-consuming.In this paper, a novel approach was developed for real-time monitoring of DNA ligation process.By using this approach, DNA ligation process can be monitored in real-time providing abundant, successive, dynamic and real-time data, which is of great importance for the mechanistic studies and dynamics research of DNA ligation.Ulteriorly, a qualification assay method for the ligase is developed, with a linear range from 4.0×10-6to 2.0×10-4U/μLand a detection limit of 4.0×10-6U/μL.

Key words: E.coli DNA ligase, DNA ligation, Molecular beacon, Real-time monitoring

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