数字农科院2.0

A sensitive one-pot ROA assay for rapid miRNA detection

文献类型： 外文期刊

作者： Hou, Zhihao;Deng, Wenpeng;Li, Alun;Zhang, Ya;Chang, Jianye;Guan, Xinyue;Chang, Yuxiao;Wang, Kaile;Wang, Xinjie;Ruan, Jue

作者机构：

关键词： （1-1-1）miRNA;Rolling circle amplification (RCA);Sensitive detection;Fluorescence detection

期刊名称： ABIOTECH

ISSN： 2096-6326

年卷期： 2024 年

页码：

收录情况： EI（2024版） ; ; CSCD（2023-2024年度）

摘要： MicroRNAs (miRNAs) and short RNA fragments (18-25 nt) are crucial biomarkers in biological research and disease diagnostics. However, their accurate and rapid detection remains a challenge, largely due to their low abundance, short length, and sequence similarities. In this study, we report on a highly sensitive, one-step RNA O-circle amplification (ROA) assay for rapid and accurate miRNA detection. The ROA assay commences with the hybridization of a circular probe with the test RNA, followed by a linear rolling circle amplification (RCA) using dUTP. This amplification process is facilitated by U-nick reactions, which lead to an exponential amplification for readout. Under optimized conditions, assays can be completed within an hour, producing an amplification yield up to the microgram level, with a detection limit as low as 0.15 fmol (6 pM). Notably, the ROA assay requires only one step, and the results can be easily read visually, making it user-friendly. This ROA assay has proven effective in detecting various miRNAs and phage ssRNA. Overall, the ROA assay offers a user-friendly, rapid, and accurate solution for miRNA detection.

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