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Entropy-Driven Catalytic G-Quadruple Cycle Amplification Integrated with Ligases for Label-Free Detection of Single Nucleotide Polymorphisms

化学 荧光 核酸 滚动圆复制 生物物理学 组合化学 DNA 生物化学 聚合酶 生物 物理 量子力学
作者
Yunshan Zhang,Fang Yang,Tuo Huang,Shijie Xu,Jing Ye,Lin Weng,Ye Hu,Haowen Huang,Shuang Li,Diming Zhang
出处
期刊:Analytical Chemistry [American Chemical Society]
被引量:2
标识
DOI:10.1021/acs.analchem.4c03057
摘要

G-Quadruplex/thioflavin (G4/THT) has become a very promising label-free fluorescent luminescent element for nucleic acid detection due to its good programmability and compatibility. However, the weak fluorescence efficiency of single-molecule G4/THT limits its potential applications. Here, we developed an entropy-driven catalytic (EDC) G4 (EDC-G4) cycle amplification technology as a universal label-free signal amplification and output system by properly programming classical EDC and G4 backbone sequences, preintegrated ligase chain reaction (LCR) for label-free sensitive detection of single nucleotide polymorphisms (SNPs). First, the positive strand LCR enabled specific transduction and preliminary signal amplification from single-base mutation information to single-strand information. Subsequently, the EDC-G4 cycle amplification reaction was activated, accompanied by the production of a large number of G4/THT luminophores to output fluorescent signals. The EDC-G4 system was proposed to address the weak fluorescence of G4/THT and obtain a label-free fluorescence signal amplification. The dual-signal amplification effect enabled the LCR-EDC-G4 detection system to accurately detect mutant target (MT) at concentrations as low as 22.39 fM and specifically identify 0.01% MT in a mixed detection pool. Moreover, the LCR-EDC-G4 system was further demonstrated for its potential application in real biological samples. Therefore, this study not only contributes ideas for the development of label-free fluorescent biosensing strategies but also provides a high-performance and practical SNP detection tool in parallel.
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