Genome-Wide Location Analyses of N6-Methyladenosine Modifications (m⁶A-Seq)

N⁶-methyladenosine-sequencing (m⁶A-seq) is a critical tool to obtain an unbiased genome-wide picture of m⁶A sites of modification at high resolution. It allows the study of the impact of various perturbations on m⁶A modification distribution and the study of m⁶A functions. Herein, we describe the m⁶A-seq protocol, which entails RNA immunoprecipitation (RIP) performed on fragmented poly(A) RNA utilizing anti-m⁶A antibodies. The captured/enriched m⁶A positive RNA fragments are subsequently sequenced by RNA-seq in parallel with background control non-immunoprecipitated input RNA fragments. Analyses reveal peaks of m⁶A enrichment containing sites of modifications analogous to chromatin modification immunoprecipitation experiments.