高等学校化学学报

• 研究论文 •    

基于链替代信号放大灵敏检测端粒酶活性的电化学方法

马艳蓉1, 江胜男2, 金燕2()   

  1. 1.北方民族大学预科教育学院, 银川 750021
    2.陕西师范大学化学化工学院, 陕西省生命分析重点实验室, 西安 710062
  • 收稿日期:2020-07-07
  • 通讯作者: 金燕 E-mail:jinyan@snnu.edu.cn
  • 基金资助:
    国家自然科学基金(21974083);中央高校基本科研业务费专项资金项目(202002002)

Sensitive and Electrochemical Detection of Telomerase Activity Based on the Signal Amplification of Strand Displacement Reaction

MA Yanrong1, JIANG Shengnan2, JIN Yan2()   

  1. 1.School of Preparatory Education,North Minzu University,Yinchuan 750021,China
    2.Analytical Chemistry for Life Science of Shaanxi Province,School of Chemistry and Chemical Engineering,Shaanxi Normal University,Xi’an 710062,China
  • Received:2020-07-07
  • Contact: JIN Yan E-mail:jinyan@snnu.edu.cn
  • Supported by:
    Supported by the National Natural Science Foundation of China(21974083);the Fundamental Research Funds for the Central Universities, China(GK 202002002)

摘要:

利用标记二茂铁基团的DNA(T-DNA)分子作为信号探针, 基于端粒酶特异性延长其底物链(TS)所引发的链替代反应, 建立了一种检测端粒酶活性的电化学信号放大法. 将巯基化的发夹型DNA分子(H-DNA)通过金-硫键自组装于金电极表面, 辅助DNA(A-DNA)与二茂铁修饰的T-DNA部分互补杂交形成双链AT-DNA; 当端粒酶存在时, 可在TS的3′末端合成TTAGGG的重复序列; A-DNA与TS延长链杂交置换出T-DNA; T-DNA与发夹H-DNA杂交使得二茂铁靠近电极表面; 一条TS延长链可以释放出多条T-DNA, 将二茂铁富集到金电极表面, 从而实现信号放大检测端粒酶活性. HeLa细胞个数在5~100范围内与电流值成正比, 最低可检测5个HeLa细胞中端粒酶的活性. 因此, 本文建立了一种简单灵敏检测端粒酶活性的电化学方法.

关键词: 端粒酶, 链替代反应, 电化学

Abstract:

An electrochemical method was developed for sensitive detection of telomerase activity based on strand displacement reaction. A ferrocene(Fc)-labeled DNA probe(T-DNA) was designed as signal probe. The thiolated hairpin DNA(H-DNA) was used as capture probe and was self-assembled on the Au electrode surface via Au-S bond. Assisted DNA(A-DNA) partially hybridized with T-DNA to form AT-DNA duplex. In the presence of telomerase, telomerase substrate(TS) primer was elongated by adding multiple TTAGGG repeats at its 3′ terminal. The elongated DNA hybridized with A-DNA to displace T-DNA which can hybridize with H-DNA. As a result, the Fc tags are close to the Au electrode surface, producing an electrochemical signal. One elongated DNA can displace multiple T-DNAs, leading to the enrichment of Fc onto the Au electrode for signal amplification. Based on this, the peak current is linearly related with the number of HeLa cells in the range of 5―100 cells, and detection limit is down to 5 HeLa cells. Therefore, it provides a simple and sensitive approach for detecting telomerase activity.

Key words: Telomerase, Strand displacement reaction, Electrochemistry

中图分类号: