Faculty Opinions recommendation of DART-seq: an antibody-free method for global m6A detection.

Abstract

m 6 A is a widespread RNA modification which influences nearly every aspect of the mRNA life cycle. Our understanding of m 6 A has been facilitated by the development of global m 6 A mapping methods, which use antibodies to immunoprecipitate methylated RNA. However, these methods have several limitations, including high input RNA requirements and cross-reactivity to other RNA modifications. Here, we present DART-Seq (deamination adjacent to RNA modification targets), an antibody-free method for detecting m 6 A sites. In DART-Seq, the cytidine deaminase APOBEC1 is fused to the m 6 A-binding YTH domain. APOBEC1-YTH expression in cells induces C to U deamination at sites adjacent to m 6 A residues, which are detected using standard RNA-Seq. DART-Seq identifies thousands of m 6 A sites in cells from as little as 10 nanograms of total RNA and can detect m 6 A accumulation in cells over time. Additionally, we use long-read DART-Seq to gain new insights into m 6 A distribution along the length of individual transcripts.