Accurate imaging of m6A on specific mRNA with proximity ligation-triggered RCA-FISH (PLTRF-m6A).

N6-methyladenosine (m6A) is one of the main internal modifications in mammalian RNA, playing an important role in gene expression and regulation. Characterization of the expression level and distribution of m6A on specific RNA in situ is necessary to understand methyladenosine-related diseases. Herein, we present a proximity ligation-triggered RCA-FISH for imaging of m6A on specific RNA (PLTRF-m6A). This method can achieve the accurate imaging of mRNA-m6A at single-molecule resolution by avoiding the extra probe-caused nonspecific amplification. And we demonstrate the efficacy of PLTRF-m6A on different breast cell lines. We envision that this approach will serve as a tool for obtaining information on methyladenosine from clinical samples.