Electrochemical immunosensor for N6-methyladenosine RNA modification detection

N6-methyladenosine (m6A) is a ubiquitous modification in mRNA and other RNAs across most eukaryotes, which is likely dynamically regulated throughout developmental processes. In this work, an electrochemical immunosensor was fabricated for RNA methylation detection using gold nanoparticles modified glassy carbon electrode (AuNPs/GCE) as substrate electrode. The assembly of 4-aminophenylboronic acid on AuNPs/GCE triggered the further immobilization of anti-m6A antibody on the substrate electrode through the specific interaction between boric acid and glycosyl of antibody, which led to the following capture of methylated RNA through the immunoreaction. A decreased electrochemical reduction signal of the redox probe of [Fe(CN)6]3− was obtained based on the synergistic effect of electrostatic repulsion and steric hindrance. The developed method showed a wide linear range for methylated RNA from 0.01 to 10 nM and the detection limit was 2.57 pM (S/N = 3). Moreover, the down-regulated expression of methylation level of mRNA in the leaves of rice seedlings after incubation with 6-benzylaminopurine was confirmed by the fabricated biosensor, indicating the great potential for assaying the expression level of methylated RNA in real sample.