A strong "on-off" electrochemical biosensor for detecting dicer-like molecules in Fusarium oxysporum f. sp. cubense race 4 (Foc4)

Banana Fusarium wilt is a devastating disease that threatens the global banana industry. The Dicer-like gene (DCL) in race 4 (Foc4) is a key pathogenic factor responsible for the disease. Sensitive and specific detection of DCL is critical in preventing the spread of Foc4 disease. Here, we report the development of a novel electrochemical adapted sensor for the detection of DCL using a ssDNA-hDNA coupled system with a dual signal amplification strategy. The sensor relies on the binding of methylene blue (MB) to both ends of a hairpin probe (hDNA), which is connected to ssDNA through hybridization-mediated, and is linked to the electrode surface via Au-S bonds. The coupling of ssDNA and hDNA with two MBs on each hairpin probe leads to dual signal amplification strategy. In the presence of DCL, hDNA carrying two MBs dissociates from ssDNA, leading to signal attenuation. Compared to sensors using ordinary hairpins, our sensor exhibits larger electrical signals and lower background currents, resulting in a wider detection range (0.01-50 nM) and lower detection limit (12 pM) for DCL. We applied this sensor to the analysis of soil, banana leaves, and fruit samples, and found it to be highly specific and stable. Our method provides a new approach for the detection of banana fusarium wilt and could be useful in assisting with its detection.