Nanopore direct RNA sequencing maps the complexity of Arabidopsis mRNA processing and m<sup>6</sup>A modification.
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Understanding genome organization and gene regulation requires insight into RNA transcription, processing and modification. We adapted nanopore direct RNA sequencing to examine RNA from a wild-type accession of the model plant Arabidopsis thaliana and a mutant defective in mRNA methylation (m 6 A). Here we show that m 6 A can be mapped in full-length mRNAs transcriptome-wide and reveal the combinatorial diversity of cap-associated transcription start sites, splicing events, poly(A) site choice and poly(A) tail length. Loss of m 6 A from 3' untranslated regions is associated with decreased relative transcript abundance and defective RNA 3' end formation. A functional consequence of disrupted m 6 A is a lengthening of the circadian period. We conclude that nanopore direct RNA sequencing can reveal the complexity of mRNA processing and modification in full-length single molecule reads. These findings can refine Arabidopsis genome annotation. Further, applying this approach to less well-studied species could transform our understanding of what their genomes encode.
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