ArticleInternational journal of molecular sciences2025
A Dual-Cycle Isothermal Amplification Method for microRNA Detection: Combination of a Duplex-Specific Nuclease Enzyme-Driven DNA Walker with Improved Catalytic Hairpin Assembly.
Article in International journal of molecular sciences, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 2 papers.
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Who cites it
2 citing papers in PubMed.
- Isolation of Exosomes from MDA-MB-231 Cells Using a Paddle Screw System and Detection of TNBC-Associated Exosomal miRNAs.Micromachines · 2026Article
- Advances in Research on Isothermal Signal Amplification Mediated MicroRNA Detection of Clinical Samples: Application to Disease Diagnosis.Biosensors · 2025Review
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Authors and funding
7 authors.
Funding
Abstract
The association between microRNAs and various diseases, especially cancer, has been established in recent years, indicating that miRNAs can potentially serve as biomarkers for these diseases. Determining miRNA concentrations in biological samples is crucial for disease diagnosis. Nevertheless, the stem-loop reverse transcription quantitative PCR method, the gold standard for detecting miRNA, has great challenges in terms of high costs and enzyme limitations when applied to clinical biological samples. In this study, an isothermal signal amplification method based on a duplex-specific nuclease (DSN) enzyme-driven DNA walker and an improved catalytic hairpin assembly (CHA) was designed for miRNA detection. First, biotin-triethylene glycol-modified trigger-releasable DNA probes were conjugated to the streptavidin-coated magnetic beads for recognizing the target miRNA. The DSN enzyme specifically hydrolyzes DNA strands when the DNA probe hybridizes with the targeted miRNA. This recycling process converts the input miRNA into short trigger fragments (catalysts). Finally, three hairpins of improved CHA are driven by this catalyst, resulting in the three-armed CHA products and a fluorescence signal as the output. This dual-cycle biosensor shows a good linear relationship in the detection of miR-21 and miR-141 over the final concentration range of 250 fM to 50 nM, presenting an excellent limit of detection (2.95 amol). This system was used to detect miR-21 and miR-141 in MCF-7 and 22RV1 cells, as well as in 1% human serum. This system can be used to evaluate the expression levels of miRNAs in different biological matrices for the clinical diagnosis and prognosis of different cancers.
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