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.　©　2024　Elsevier　B.V.