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A critical appraisal of base-resolution m6A profiling techniques.

Natalia Wawrzyniak·Eduardo Eyras·Justin J-L Wong
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Abstract

N6-methyladenosine (m6A) is the most prevalent internal modification in eukaryotic mRNA, influencing RNA fate and gene regulation. Early antibody-based approaches enabled transcriptome-wide profiling but lacked resolution and quantitative accuracy. Newer approaches now achieve base-resolution m6A detection using improved crosslinking, chemical or enzymatic conversion, and single-molecule sequencing. Antibody-free methods provide quantitative stoichiometry from minimal input, while nanopore direct RNA sequencing offers real-time, single-molecule readouts across entire transcriptomes. Collectively, these methods form a versatile toolkit that integrates global mapping with precise site-level analysis, advancing knowledge of context-dependent m6A regulation in physiology and disease. This review compares their principles, strengths, and limitations to guide method selection and highlight how next-generation epitranscriptomic tools are paving the way for clinical and therapeutic applications.

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HumansRNA MethylationAdenosineEpitranscriptomeEpitranscriptomicsAnimalsRNA, Messengerantibody-based approachesepitranscriptomicsm(6)A RNA modificationnanopore sequencingsingle-nucleotide RNA methylation mappingJournal ArticleReview

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Natalia Wawrzyniak, Eduardo Eyras, Justin J-L Wong. 2026-07-16. A critical appraisal of base-resolution m6A profiling techniques.. https://doi.org/10.1016/j.tibs.2026.06.010

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