The Study Of Electrochemical Biosensing For Detection Of MicroRNA Based On Nucleic Acid Amplification Technology

MicroRNAs(miRNAs)belong to a class of endogenous,non-coding small RNA molecules that regulate the expression of gene.The abnormal expression of mi RNA is associated with many diseases,especially tumors,making miRNA as a new valuable biomarker in early clinical diagnosis.Therefore,it is great of significance to develop a simple,rapid and highly sensitive method for the detection of micro RNA.Electrochemical biosensor has the advantages of low cost,simple operation,fast response,and high sensitivity,which has been widely used in the detection of biomolecules recent years.Meanwhile,in order to further improve the sensitivity of miRNA detection,different nucleic acid signal amplification strategies have been widely developed in the field of biosensing.In this study,a simple,rapid and sensitive electrochemical biosensor was developed for detection of miRNA by integrating strand displacement amplification(SDA)and catalytic hairpin assembly(CHA)techniques.In the presence of target miRNA,SDA was initiated to obtain a large number of DNA priming strands associated with the target miRNA,which catalyzes the self-assembly of the hairpins and product a large amount of biotin-labeled double-stranded DNA.And then,the obtained biotin-labeled dsDNA was immobilized on the surface of the electrode modified with auxiliary probe.Later,ST-AP was modified onto the electrode surface by the specific recognition of biotin and streptavidin.When the ?-NPP was in the test buffer,?-NPP was catalyzed by ST-AP to in situ produce ?-NP on the surface of the electrode.Finally,the electrochemical signal of ?-NP was correlated with the concentration of target miRNA to achieve high sensitivity detection.Under the optimal experimental conditions,the proposed biosensor had a linear range from 0.1 pM to 10 nM,and the determination of target miRNA was down to 60 fM,and the single based mismatched sequence could be clearly distinguished.Therefore,the constructed biosensor will provide new technical support for the clinical detection of miRNA.