Detection of m6A RNA modifications at single-nucleotide resolution using m6A-selective allyl chemical labeling and sequencing

As the most abundant internal mRNA modification, N6-methyladenosine (m6A) was involved in almost all the aspects of RNA metabolism. Here, we introduce our protocol for m6A-SAC-seq, which enables the whole transcriptome-wide mapping of m6A RNA modification at single-nucleotide resolution with stoichiometry information. m6A-SAC-seq relies on selective allyl labeling of m6A by specific methyltransferase and chemical treatment that introduce mutation upon reverse transcription. The technique only requires ∼30 ng of input RNA. For complete details on the use and execution of this protocol, please refer to Hu et al. (2022). Keywords: Bioinformatics; Biotechnology and bioengineering; Chemistry; Genomics; Molecular biology; Molecular/Chemical probes; Protein biochemistry; Protein expression and purification; Sequence analysis.