Protocol for antibody-based m6A sequencing of human postmortem brain tissues

Here, we present an optimized N6-methyladenosine (m6A) immunoprecipitation sequencing for human postmortem brain tissue, building on previously established approaches. We describe steps for RNA extraction, RNA shearing, RNA immunoprecipitation, and library preparation. The protocol includes optimization data for each step, such as m6A profiles from total RNA versus poly(A) RNA as an input, a comparison of different commercial m6A antibodies, and immunoprecipitation with varying RNA/antibody ratios. This protocol allows reliable capture of m6A peaks from human postmortem brain tissue. For complete details on the use and execution of this protocol, please refer to Mitsuhashi et al.¹.