Signal-On Fluorescence Biosensor for Highly Sensitive Detection of miRNA-21 Based on DNAzyme Assisted Double-Hairpin Molecular Beacon

Abstract

Although miRNAs exist in small quantities in the human body, they are closely related to the abnormal expression of genes in diseases such as tumors. Therefore, sensitive detection of miRNAs is very important for the prevention and treatment of various tumors and major diseases. The purpose of this study is to develop a label-free sensing strategy based on the co-action of double-hairpin molecular beacons and deoxyribozymes (DNAzymes) for highly sensitive detection of miRNA-21. The target miRNA-21 promotes the assembly of DNAzyme with a complete catalytic core region. At the presence of Mg2+, DNAzyme cuts a substrate into short chains, which open the double hairpin molecular beacon, and then form G-quadruplexs at both ends, specifically binding more ThT to generate a amplified fluorescent signal. The cut substrate will be replaced by the uncut ones in the next stage, increasing the concentration of reactants, and thus further improving the fluorescence intensity. This DNAzyme assisted double hairpin molecular beacon has a certain degree of discrimination for substances with single base mismatches, and the detection limit of miRNA-21 is 0.13 pM, lower than that of the many other analysis. Further, this detection has good selectivity and sensitivity in serum. Therefore, this strategy provides a simple, fast and low-cost platform for the sensitive detection of miRNA-21, having potential applications in early cancer diagnosis.

Nanosensor Based on the Dual-Entropy-Driven Modulation Strategy for Intracellular Detection of MicroRNA

“…Until now, a variety of biosensors have been successfully developed for detection of miRNA with their advantages of good sensitivity, accuracy, and ease of miniaturization. − However, due to the low abundance of miRNA, the discrimination of the change of their concentration has certain challenges. Therefore, various nucleic acid-based signal amplification strategies have been developed to further improve the sensitivity of biosensors. , Among them, polymerase-dependent amplification technology relies on the exponential growth of nucleic acid under the catalysis of polymerase, which can significantly improve the sensitivity. − However, once the reaction system is polluted, it is easy to have false positive results. , Polymerase-independent amplification technology can effectively amplify the detection signal while reduce the cost and time of a single detection, which is of great significance for the development of miRNA detection schemes. , Hybrid chain reaction (HCR) and catalytic hairpin assembly (CHA) are frequently used enzyme-free amplification technologies; yet, they require expertise in precise sequence design.…”