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Updated: Sep 2, 2025
Probe-based Real-time PCR Approaches for Quantitative Measurement of microRNAs
Published on: April 14, 2015
Fluorescence energy transfer biosensing platform based on hyperbranched rolling circle amplification and multi-site
Hongkun Li1, Qianqian Cai1, Di Wu1
1Key Laboratory of Optic-electric Sensing and Analytical Chemistry for Life Science, MOE, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao, 266042, PR China.
Abstract:
A novel fluorescence biosensor based on hyperbranched rolling circle amplication (HRCA) and multi-site strand displacement amplification (SDA) strategy was constructed to realize super-sensitive detection of miRNA-21. The target miRNA-21 was specifically used to trigger HRCA reaction and generated abundant DNA sequences with different lengths. These sequences could initiate the SDA reaction with hairpin HP1 and HP2 to form the fluorescence signal double-stranded bodies (FSDB), and produced the fluorescence resonance energy transfer (FRET) for target assay. The circle template, reverse primer, hairpin HP1 and HP2 were skillfully designed to much improve the assay selectivity. This FRET signal ratio-based strategy not only avoided the false positive signal produced by traditional detection methods, but also minimized the influence of system fluctuation. So the detection limit of the target could reach fM level. In addition, this method could also be applied to the detection of other miRNAs, proteins and biomolecules, and had great potential in biomedical research, environmental detection and clinical diagnostic applications.
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