Ultrasensitive miRNA Detection: A Novel DNA Nanomachine Using Split-type Molecular Beacons-Mediated Cascade Amplification for Cancer Diagnostics

MicroRNAs (miRNAs) are crucial regulators in various pathological and physiological processes, and their misregulation is a hallmark of many diseases. In this study, we introduce an advanced DNA nanomachine using split-type molecular beacons (STMBs) for sensitive detection of miR-21, a key biomarker in cancer diagnostics. Utilizing an innovative STMB-mediated cascade strand displacement amplification (STMB-CSDA) technique, our approach offers a powerful means for the precise quantification of miRNAs, using miR-21 as a primary example. The system operates through target-induced linkage of STMBs, initiating a series of strand displacement amplifications resulting in exponential signal amplification. Coupled with the precision of T4 DNA ligase, this mechanism translates minimal miRNA presence into significant fluorescence signals, offering detection sensitivity as low as 5.96 pM and a dynamic range spanning five orders of magnitude. Characterized by its high specificity, which includes the ability to identify single-base mismatches, along with its user-friendly design, our method represents a significant leap forward in miRNA analysis and molecular diagnostics. Its successful application in examining total RNA from cancer cells and clinical serum samples demonstrates its immense potential as a groundbreaking tool for early cancer detection and gene expression studies, paving the way for the next generation of non-invasive diagnostics in personalized healthcare.