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Three-Way Junction-Assisted Rolling Circle Amplification Integrated with trans-Cleavage of Cas12a for Sensitive and Reliable Detection of miRNA

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Abstract

MicroRNAs (miRNAs) serve a crucial role in numerous biological processes, such as acute pancreatitis development. Due to its low abundance and high similarity among homogeneous family members, sensitive and reliable detection of microRNA remains a formidable challenge. By combining the three-way junction-assisted rolling circle amplification (RCA) with the trans-cleavage of Cas12a, we propose a novel fluorescent technique for sensitive miRNA detection. In order to increase the amplification efficiency of RCA-based methods, catalytic hairpin amplification (CHA) is incorporated into the RCA process, playing the roles of specific target recognition and three-way junction formation. Consequently, the method demonstrated a six-orders-of-magnitude detection range and a LOD as low as 27 aM, making it a promising method for the early diagnosis of various diseases.

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All data generated and analyzed during this study are included in this article.

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Acknowledgements

We appreciate the financial support from the Chongqing Medical University. We also thank the director of central laboratory for providing essential equipment.

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Authors and Affiliations

  1. Emergency and Critical Care Center, The Third Affiliated Hospital of Chongqing Medical University, Chongqing City, 401120, China

    Zhiquan Wu

  2. Department of Hematology, The Third Affiliated Hospital of Chongqing Medical University, Shuanghu Branch Road, Yubei District, Chongqing City, 401120, China

    Shuqi Zhao

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  1. Zhiquan Wu
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Shuqi Zhao designed and wrote the manuscript; Zhiquan Wu performed experiments and analyzed obtained data.

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Correspondence to Shuqi Zhao.

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Wu, Z., Zhao, S. Three-Way Junction-Assisted Rolling Circle Amplification Integrated with trans-Cleavage of Cas12a for Sensitive and Reliable Detection of miRNA. Appl Biochem Biotechnol 196, 3115–3125 (2024). https://doi.org/10.1007/s12010-023-04691-8

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  • DOI: https://doi.org/10.1007/s12010-023-04691-8

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