A Binary DNAzyme Machine (BDM) for Selective Amplification-Free Detection of microRNA

Objective: Develop a PCR-free DNA nanomachine (BDM) for selective picomolar detection of plasma miR-371a. Methods: A split 10–23 deoxyribozyme was engineered with substrate-delivering hooks and mismatch-introducing arms. The system was validated at 35°C using synthetic targets and patient plasma. Results and Discussion: BDM achieved a 1.6 pM limit of detection, showing a 9-fold sensitivity increase over hook-less controls. It maintained excellent specificity (LOD 1.3 pM) against a 25-fold excess of homologs and successfully identified clinical seminoma samples. Conclusions: BDM offers a sensitive, contamination-resistant alternative to RT-qPCR for non-invasive cancer liquid biopsies.