Bifunctional Y-shaped probe combined with dual amplification for colorimetric sensing and molecular logic operation of two miRNAs

The abnormal expression of miRNA is closely related to various human diseases. In particular, the sensitivity detection of miRNA expression level is of great significance for the early diagnosis and prognosis of cancer. In this paper, we designed a Y-shaped DNA probe, using miRNA-21 and miRNA-141 as the dual input signals of AND logic gate. By combining with EXO III assisted target recycle and DNA hybridization chain reaction (HCR), we have realized dual signal amplification for detection of two miRNAs. In short, the Y-shaped DNA probe consists of two parts: the miRNA target binding region and the HCR initiator. When the two miRNAs are present at the same time, the target binding region specifically recognizes the target to generate two circulators, and then the HCR initiator is released. The EXO III specific cleavage two circulator, and release the target again which achieves the first step of signal amplification. After that, HCR was started by the split initiator generated in the first stage of continuous cycle, and the second step of signal amplification was realized. Thanks to the sensitive color change of gold nanoparticles in response to salt, we achieved ultra-high sensitivity for visual detection of miRNA-21 and miRNA-141. Under optimal conditions, the detection limit of both miRNA is 3 pM and the linear range is 10 pM to 0.4 nM. The method we designed could be applied in early detection and diagnosis of cancer.