基于无细胞 RNA 转录系统靶向转化和双链特异性核酸酶辅助循环扩增的抗坏血酸电化学发光生物传感器

IF 6.7 1区 化学 Q1 CHEMISTRY, ANALYTICAL Analytical Chemistry Pub Date : 2024-10-27 DOI:10.1021/acs.analchem.4c0420610.1021/acs.analchem.4c04206
Ming Chen, Hongfu Zeng, Fang Luo, Yunjian Huang*, Cuiying Lin*, Jian Wang, Bin Qiu and Zhenyu Lin*, 
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引用次数: 0

摘要

首次将无细胞 RNA 转录系统与电化学发光(ECL)检测技术相结合,开发出一种抗坏血酸(AA,作为模型目标)生物传感器。该生物传感器由分别用炔基和叠氮基修饰的单链 DNA(ssDNA)序列和一个不完整的基因电路框架组成。加入目标 AA 和铜离子后,两个 ssDNA 序列将通过点击化学反应连接起来。这样就能重建完整的基因回路。用二茂铁(Fc)标记的 ssDNA(Fc-DNA)首先被固定在三(2,2′-联吡啶)氯化钌(II)六水合物掺杂的二氧化硅纳米粒子(Ru@SiO2 NPs)修饰电极上。由于 Fc 对 Ru@SiO2 的淬灭作用,导致检测到的 ECL 较低。转录的 RNA 序列辅助双链特异性核酸酶(DSN)切割 ssDNA-Fc,从而增强了系统的 ECL。最佳实验条件显示,ECL 信号与 AA 的对数浓度呈线性相关,检测范围从 100 nM 到 1 mM,检测限为 45 nM。这一创新方法拓展了无细胞 RNA 转录系统在生物传感领域的应用。
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Electrochemiluminescence Biosensor for Ascorbic Acid Based on Target Transformation of Cell-Free RNA Transcription System and Duplex-Specific Nuclease-Assisted Recycling Amplification

A cell-free RNA transcription system had been coupled with electrochemiluminescence (ECL) detection technology for the first time to develop an ascorbic acid (AA, acting as a model target) biosensor. The biosensor is composed of single-stranded DNA (ssDNA) sequences modified with alkynyl and azido groups, respectively, alongside an incomplete gene circuit framework. The addition of target AA and copper ions will cause the linkage of the two ssDNA sequences through a click chemistry reaction. This results in the subsequent reconstruction of a complete gene circuit. The reconstituted gene circuit, in conjunction with the T7 RNA polymerase, drives the transcription of substantial quantities of RNA. ssDNA labeled with ferrocene (Fc) (Fc-DNA) had been immobilized on a tris(2,2′-bipyridyl) ruthenium(II) chloride hexahydrate-doped SiO2 nanoparticle (Ru@SiO2 NPs) modified electrode first. The quenching effect of Fc on Ru@SiO2 causes the low ECL detected. The transcribed RNA sequence assisted double-stranded specific nuclease (DSN) to cut the ssDNA-Fc and the ECL of the system was enhanced. Optimal experimental conditions reveal that the ECL signal exhibits a linear correlation with the logarithmic concentration of AA, spanning a detection range from 100 nM to 1 mM, with a detection limit of 45 nM. This innovative methodology expands the utility of a cell-free RNA transcription system within the realm of biosensing applications.

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来源期刊
Analytical Chemistry
Analytical Chemistry 化学-分析化学
CiteScore
12.10
自引率
12.20%
发文量
1949
审稿时长
1.4 months
期刊介绍: Analytical Chemistry, a peer-reviewed research journal, focuses on disseminating new and original knowledge across all branches of analytical chemistry. Fundamental articles may explore general principles of chemical measurement science and need not directly address existing or potential analytical methodology. They can be entirely theoretical or report experimental results. Contributions may cover various phases of analytical operations, including sampling, bioanalysis, electrochemistry, mass spectrometry, microscale and nanoscale systems, environmental analysis, separations, spectroscopy, chemical reactions and selectivity, instrumentation, imaging, surface analysis, and data processing. Papers discussing known analytical methods should present a significant, original application of the method, a notable improvement, or results on an important analyte.
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