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Theranostics 2018; 8(10):2646-2656. doi:10.7150/thno.24480 This issue Cite

Research Paper

Catalytic hairpin assembly gel assay for multiple and sensitive microRNA detection

Wenhao Dai¹, Jing Zhang², Xiangdan Meng¹, Jie He³, Kai Zhang¹, Yu Cao¹, Dongdong Wang¹, Haifeng Dong^1,✉, Xueji Zhang^1,✉

1. Research Center for Bioengineering and Sensing Technology, Beijing Key Lab for Bioengineering and Sensing Technology, School of Chemistry and bioengineering, University of Science & Technology Beijing, Beijing 100083, P.R. China.
2. School of Petrochemical Engineering, School of Food Science and Technology, Changzhou University, Changzhou 213164, P.R. China.
3. School of Computer and Communication Engineering, University of Science & Technology Beijing, Beijing 100083, P.R. China.

✉ Corresponding author: hfdongedu.cn; zhangxuejiedu.cn. Phone/Fax: +86 010 82376576.

Abstract

As important modulators of gene expression, microRNAs (miRNAs) have been identified as promising biomarkers with powerful predictive value in diagnosis and prognosis for several diseases, especially for cancers. Here we report a facile, multiple and sensitive miRNA detection method that uses conventional gel electrophoresis and catalytic hairpin assembly (CHA) system without any complex nanomaterials or enzymatic amplification.

Methods: In this study, three pairs of hairpin probes are rationally designed with thermodynamically and kinetically preferable feasibility for the CHA process. In the present of target miRNA, the stem of the corresponding hairpin detection probe (HDP) will be unfolded and expose the concealed domain. The corresponding hairpin assistant probe (HAP) then replaces the hybridized target miRNA to form specific HDP/HAP complexes and releases miRNA based on thermodynamically driven entropy gain process, and the released miRNA triggers the next recycle to produce tremendous corresponding HDP/HAP complexes.

Results: The results showed that the CHA gel assay can detect miRNA at fM levels and shows good capability of discriminating miRNA family members and base-mismatched miRNAs. It is able to analyze miRNAs extracted from cell lysates, which are consistent with the results of conventional polymerase chain reaction (PCR) method. Depending on the length of the designed hairpin probes, the CHA gel assay consisting of different hairpin probes effectively discriminated and simultaneously detected multiple miRNAs in homogenous solution and miRNAs extracted from cell lysates.

Conclusion: The work highlights the practical use of a conventional gel electrophoresis for sensitive interesting nucleic acid sequences detection.

Keywords: microRNA, multiple microRNA detection, catalytic hairpin assembly, gel electrophoresis, PCR

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