Electrochemical detection of circRNAs based on the combination of back-splice junction and duplex-specific nuclease

Covalently closed circular RNA (circRNA) has potential implications for disease diagnosis and therapy. However, current methods for circRNA detection are infrequent, and require RNase R pretreatment in general, which is deviation-oriented and labor-intensive. Herein, we have developed a simple electrochemical method for direct detection of circRNA. Firstly, the hairpin probes are designed to identify circRNA based on back-splice junction (BSJ) sites. Then, the selection of substrate length by duplex-specific nuclease (DSN) is utilized to trigger DSN-assisted amplification, resulting in significantly amplified signal. Therefore, the limit of detection of the proposed method for circRNA can reach as low as 3.47 fM, which may contribute to a better selectivity, reproducibility, and stability. This method has also been employed to assay circRNA in human serum samples, which may hold great potential in the clinical diagnosis applications.