无酶催化发夹自组装用于动力学竞争适配体传感策略灵敏检测蓖麻毒素

引用本文: 齐丽娟, 杨媚婷, 杜衍. 无酶催化发夹自组装用于动力学竞争适配体传感策略灵敏检测蓖麻毒素[J]. 分析化学, 2021, 49(6): 1025-1034. doi: 10.19756/j.issn.0253-3820.211053 shu

Citation: QI Li-Juan, YANG Mei-Ting, DU Yan. Enzyme-Free Catalytic Hairpin Assembly Utilized in A Kinetically Competitive Aptasensor for Sensitive Detection of Ricin Toxin A Chain[J]. Chinese Journal of Analytical Chemistry, 2021, 49(6): 1025-1034. doi: 10.19756/j.issn.0253-3820.211053 shu

无酶催化发夹自组装用于动力学竞争适配体传感策略灵敏检测蓖麻毒素

齐丽娟 ^1,2, ,
杨媚婷 ^{1,
,} ,
杜衍 ^{1,2,
,}

1.
中国科学院长春应用化学研究所, 电分析化学国家重点实验室, 长春 130022;
2.
中国科学技术大学, 应用化学与工程学院, 合肥 230026

通讯作者: 杨媚婷,E-mail:duyan@ciac.ac.cn; 杜衍,E-mail:mtyang@ciac.ac.cn

基金项目:
国家自然科学基金项目（Nos.21874129，31960506）和吉林省科技厅国际科技合作项目（No.20200801044GH）资助。

摘要: 构建了一种新型的基于动力学竞争的适配体传感策略，通过无酶催化发夹自组装（CHA）反应进行信号放大和输出，实现了蓖麻毒素A链（RTA）的选择性灵敏检测。阻碍链（Blocker）能够分别与适配体（RA80）和发夹探针（HP1）竞争结合，而优先与RA80结合的RTA会影响RA80和HP1对Blocker的竞争反应，使更多的Blocker与HP1杂交。随后，与HP1结合的Blocker可通过HP1上修饰的生物素与磁珠表面链霉亲和素的共价键和作用，经过磁性作用分离出来，继而作为CHA的催化链，催化CHA产生荧光信号。随着RTA浓度增大，分离出的CHA催化链浓度增加，荧光信号增强，基于此可以建立输出荧光信号与RTA浓度的关系。为降低背景信号，在CHA中引入错配碱基对，通过减少呼吸作用或自身变构引起的非催化反应，有效改善了检测信号。本传感器对RTA的检出限为0.88 nmol/L，与文献报道的对蓖麻毒素的检出限相当，在食品（包括蔗糖、小苏打和面粉等）中RTA的回收率为91.2%~107.9%。本检测方法拓展了适配体识别机制，有效解决了基于平衡的传统互补链替换法中针对长链适配体设计阻碍链的难题，竞争元素设计简便，不需要冗长的平衡过程和适配体的修饰，为序列冗长、结构复杂的RNA适配体的应用奠定了基础。

关键词:

English

Enzyme-Free Catalytic Hairpin Assembly Utilized in A Kinetically Competitive Aptasensor for Sensitive Detection of Ricin Toxin A Chain

QI Li-Juan ^1,2, ,
YANG Mei-Ting ^{1,
,} ,
DU Yan ^{1,2,
,}

1.
State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, China;
2.
School of Applied Chemistry and Engineering, University of Science & Technology of China, Hefei 230026, China

Corresponding author: YANG Mei-Ting, ; DU Yan,

Received Date: 21 January 2021
Revised Date: 12 April 2021
Fund Project:
Supported by the National Natural Science Foundation of China (Nos.21874129, 31960506) and the International Scientific Cooperation Project of Jilin Scientific and Technological Development Program, China (No.20200801044GH).

Abstract: A novel kinetically competitive aptasensor for detection of ricin toxin A chain (RTA) was fabricated based on the catalytic hairpin assembly (CHA) for signal amplification and transduction. The elements of kinetic competition contain aptamers (RA80), hairpin probes (HP1) and blockers, wherein the blockers kinetically compete with RA80 and HP1, but pre-binding of the target to its aptamer alters this kinetic competition and leads to more blocker binding to HP1. Then, more blockers binding to HP1 were separated as the catalyst for CHA reaction and increased fluorescence intensity. Thus, an indirect correlation between the fluorescence intensity and RTA concentration was constructed. Besides, the mismatched base pair introduced in the CHA circuit impeded uncatalyzed strand exchange reactions due to breathing or the mis-folding of nucleic acids into alternative conformers, which resulted in the decline of background signal and the improvement of signal-to-noise ratio. The limit of detection (LOD) for RTA was 0.88 nmol/L, which was comparable or even superior to other RTA determination strategies. The recoveries ranging from 91.2% to 107.9% were also satisfactory for RTA analysis in food powder. The proposed approach expanded the aptamer recognition mechanism and overcame the difficulty of the traditional antisense displacement strategy, bringing a bright hope to design and screen blockers for RNA aptamer with long sequence and complicated secondary structure without necessity of aptamer modification and requirement of lengthy pre-equilibration.

Key words:

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沈阳化工大学材料科学与工程学院沈阳 110142

无酶催化发夹自组装用于动力学竞争适配体传感策略灵敏检测蓖麻毒素

通讯作者: 杨媚婷,E-mail:duyan@ciac.ac.cn; 杜衍,E-mail:mtyang@ciac.ac.cn

English

Enzyme-Free Catalytic Hairpin Assembly Utilized in A Kinetically Competitive Aptasensor for Sensitive Detection of Ricin Toxin A Chain

Corresponding author: YANG Mei-Ting, ; DU Yan,

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通讯作者: 陈斌, bchen63@163.com

中国化学会秘书处

无酶催化发夹自组装用于动力学竞争适配体传感策略灵敏检测蓖麻毒素

通讯作者: 杨媚婷,E-mail:duyan@ciac.ac.cn; 杜衍,E-mail:mtyang@ciac.ac.cn

English

Enzyme-Free Catalytic Hairpin Assembly Utilized in A Kinetically Competitive Aptasensor for Sensitive Detection of Ricin Toxin A Chain

Corresponding author: YANG Mei-Ting, ; DU Yan,

CCS Chemistry：嵌段共聚物自组装成 “三明治”二维有机材料，实现纳米级压力传感功能

CCS Chemistry：颜徐州、鲍哲南： 你的皮肤可能是一片SPMs

CCS Chemistry：工作高效不疲劳，只须让催化剂动起来

CCS Chemistry：静电组装导向聚合- 聚电解质纳米凝胶量化制备新策略

CCS Chemistry：金黄色葡萄球菌醛基脱氢酶是如何识别特异性底物的？

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通讯作者: 陈斌, bchen63@163.com

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