Development of a two-in-one integrated assay for the analysis of circRNA-microRNA interactions.

The competitive endogenous RNA hypothesis is a new mechanism of RNA dialogue, in which circRNA-miRNA interaction (cmRRI) is found to be widely involved in the regulation of gene expression in tumors and other diseases. It is urgent but challenging to develop a convenient and efficient method to study the interaction between target circRNA and the candidate miRNAs. In this work, a biosensing method that allows directly analyzing cmRRI has been developed, so as to reveal the RNA dialogue strategy. The sensing system uses a bifunctional magnetic bead for the capture of target circRNA/miRNA complex as well as the signal amplification. Based on the nature of circRNA as a miRNA sponge, only if the target circRNAs and its regulatory miRNAs coexist as a complex, can the rolling circle amplification reaction be initiated to give a fluorescent signal as the output. Compared with traditional methods where the circRNA and its regulatory miRNAs have been separately analyzed, our design allows the integrated profiling of specific cmRRI by correlation characterization of two correlative RNAs, which represents a function-oriented method. The presented method also shows the analysis of the potential binding affinity of candidate miRNAs to target circRNAs. Furthermore, we have verified the ability of the sensor to directly detect cmRRI in biological samples, which reveals the promising applicability of this method for biomedical and clinical researches in the future.