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Dual-probe fluorescent biosensor based on T7 exonuclease-assisted target recycling amplification for simultaneous sensitive detection of microRNA-21 and microRNA-155

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Abstract

Effective and simultaneous monitoring of the abnormal expression of certain microRNAs (miRNAs), especially for miRNA-21 and miRNA-155, can indicate drug resistance in lung cancer. In this work, T7 exonuclease (T7 Exo)-assisted target recycling amplification coupled with the extensive fluorescence quenching of graphene oxide (GO) was designed for the simultaneous detection of miRNA-21 and miRNA-155 using FAM- and ROX-labeled single-strand DNA probes. Through this method, the variable emission intensities of FAM and ROX caused by the introduction of miRNA-21 and miRNA-155, respectively, were obtained with high sensitivity. The method exhibited excellent analytical performance for simultaneous detection of miRNA-21 and miRNA-155 without cross-interference. The linear range was from 0.005 nM to 5 nM over three orders of magnitude, with detection limits as low as 3.2 pM and 4.5 pM for miRNA-21 and miRNA-155, respectively. Furthermore, the recovery (92.49–103.67%) and relative standard deviation (RSD < 4.8%) of the standard addition test of miRNA-21 and miRNA-155 in human plasma suggested the potential for drug resistance warning in clinical practice via this simple strategy.

Graphical abstract

A homogeneous T7 Exo-assisted signal amplification combined with GO quenching platform was developed for accurate, sensitive and simultaneous analysis of miRNA-21 and miRNA-155 for drug resistance warning in lung cancer. This simple method exhibited a wide linear range and low LODs for miR-21 and miR-155.

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Acknowledgements

The authors gratefully acknowledge the financial support of the National Natural Science Foundation of China (21705021, 21775023 and 21904019), Joint Funds for the Innovation of Science and Technology, Fujian province (2017Y9121, 2019Y9011), Open project funded by Key laboratory of biological genetic resources from Ministry of Natural Resources (HY201703) and Medical Innovation Project of Fujian Province of China (2016-CX-44).

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Author notes

  1. Yanjie Zheng and Jinyuan Chen contributed equally to this work.

Authors and Affiliations

  1. Department of Pharmaceutical Analysis, School of Pharmacy, Fujian Medical University, Fuzhou, 350122, Fujian, China

    Yanjie Zheng, You Li, Yichun Xu, Li Chen, Wei Chen, Ailin Liu, Xinhua Lin & Shaohuang Weng

  2. The Central lab, the First Affiliated Hospital of Fujian Medical University, Fuzhou, 350005, Fujian, China

    Jinyuan Chen

  3. Department of Pharmacy, Fuzhou Pulmonary Hospital of Fujian, Fuzhou, 350008, Fujian, China

    You Li

  4. Nano Biomedical Technology Research Center, Fujian Medical University, Fuzhou, 350122, Fujian, China

    Wei Chen, Ailin Liu & Xinhua Lin

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  1. Yanjie Zheng
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Correspondence to Xinhua Lin or Shaohuang Weng.

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This study conformed to the ethical guidelines of the Declaration of Helsinki and was approved by the Ethics Committee for Human Research, The First Affiliated Hospital of Fujian Medical University. Human serum samples used in this study do not have any identifying information about any of the participants, and all participants provided written informed consent.

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Zheng, Y., Chen, J., Li, Y. et al. Dual-probe fluorescent biosensor based on T7 exonuclease-assisted target recycling amplification for simultaneous sensitive detection of microRNA-21 and microRNA-155. Anal Bioanal Chem 413, 1605–1614 (2021). https://doi.org/10.1007/s00216-020-03121-6

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  • DOI: https://doi.org/10.1007/s00216-020-03121-6

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