Colorimetric And Visual Determination Of MiRNA Via Cycling Signal Amplication Using T7 Exonuclease

MicroRNAs(miRNAs)are a class of small(18-23 nucleotides),endogenous,single-stranded RNA molecules that regulate the expression of genes in many cellular processes.The unnatural expression of miRNA may not only lead to serious diseases such as cancers,nervous diseases and diabetes but also act as significant signaling molecules in disease development and progression.However,given the unique characteristics of miRNAs,including short-length,low abundance,and sequence homology among family members,sensitive detection of miRNAs has remained a great challenge.Thus,the quantitative analysis of miRNA is urgently needed as novel biomarkers.In this study,a strategy based on T7 exonuclease "signal-off" was established,and the visualization of miRNA was successfully achieved in homogeneous system.In the absence of the target miRNA,the single-stranded capture probe that can't be cleaved by the T7 exonuclease opens the hairpin structure probe containing the G4 sequence,and the absorbance reaches the maximum.In the presence of target miRNA,taking advantage of T7 exonuclease,miRNA can hybridize with capture probe to form DNA / RNA double strand and the capture probe in the double strand can be cleaved.This sequence of reaction lead to decrease the concentration of capture probe and the number of opened hairpin probe,resulting in signal reduce.Under the optimal experimental conditions,the minimum detection limit of miRNA can reach 0.6 p M(S / N = 3).The linear range is 10 p M to 100 n M.This method shows a good performance in detecting homology miRNA and complex matrix.Therefore,the sensing strategy provide a new method for the detection of miRNAs,providing technical support for the diagnosis and treatment of clinically relevant diseases.