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AlkAniline-Seq: Profiling of m7 G and m3 C RNA Modifications at Single Nucleotide Resolution.

par Marchand, Virginie;Ayadi, Lilia;Ernst, Felix ;Hertler, Jasmin;Bourguignon-Igel, Valérie;Galvanin, Adeline;Kotter, Annika;Helm, Mark;Lafontaine, Denis ;Motorin, Yuri
Référence Angewandte Chemie, 57, 51, page (16785-16790)
Publication Publié, 2018-12-01

Article révisé par les pairs

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Résumé :

RNA modifications play essential roles in gene expression regulation. Only seven out of >150 known RNA modifications are detectable transcriptome-wide by deep sequencing. Here we describe a new principle of RNAseq library preparation, which relies on a chemistry based positive enrichment of reads in the resulting libraries, and therefore leads to unprecedented signal-to-noise ratios. The proposed approach eschews conventional RNA sequencing chemistry and rather exploits the generation of abasic sites and subsequent aniline cleavage. The newly generated 5'-phosphates are used as unique entry for ligation of an adapter in library preparation. This positive selection, embodied in the AlkAniline-Seq, enables a deep sequencing-based technology for the simultaneous detection of 7-methylguanosine (m7 G) and 3-methylcytidine (m3 C) in RNA at single nucleotide resolution. As a proof-of-concept, we used AlkAniline-Seq to comprehensively validate known m7 G and m3 C sites in bacterial, yeast, and human cytoplasmic and mitochondrial tRNAs and rRNAs, as well as for identifying previously unmapped positions.