Long-read sequencing technology enables highly accurate detection of allele-specific RNA expression, providing insights into the effects of genetic variation on splicing and RNA abundance. Furthermore, the ability to directly sequence RNA using the Oxford Nanopore technology promises the detection of RNA modifications in tandem with ascertaining the allelic origin of each molecule. Here, we leverage these advantages to determine allele-biased patterns of N6-methyladenosine (m6A) modifications in native mRNA. We utilized human and mouse cells with known genetic variants to assign allelic origin of each mRNA molecule combined with a supervised machine learning model to detect read-level m6A modification ratios. Our analyses revealed the importance of sequences adjacent to the DRACH-motif in determining m6A deposition, in addition to allelic differences that directly alter the motif. Moreover, we discovered allele-specific m6A modification (ASM) events with no genetic variants in close proximity to the differentially modified nucleotide, demonstrating the unique advantage of using long reads and surpassing the capabilities of antibody-based short-read approaches. This technological advancement promises to advance our understanding of the role of genetics in determining mRNA modifications.

中文翻译：

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长读长 RNA 测序显示等位基因特异性 N6-甲基腺苷修饰


长读长测序技术能够高度准确地检测等位基因特异性 RNA 表达，从而深入了解遗传变异对剪接和 RNA 丰度的影响。此外，使用 Oxford Nanopore 技术直接对 RNA 进行测序的能力有望在确定每个分子的等位基因起点的同时检测 RNA 修饰。在这里，我们利用这些优势来确定天然 mRNA 中 N6-甲基腺苷 （m6A） 修饰的等位基因偏倚模式。我们利用具有已知遗传变异的人类和小鼠细胞来分配每个 mRNA 分子的等位基因起点，并结合监督机器学习模型来检测读取水平的 m6A 修饰率。我们的分析揭示了与 DRACH 基序相邻的序列在确定 m6A 沉积中的重要性，以及直接改变基序的等位基因差异。此外，我们发现了等位基因特异性 m6A 修饰 （ASM） 事件，在差异修饰核苷酸附近没有遗传变异，证明了使用长读长的独特优势，并超越了基于抗体的短读长方法的能力。这项技术进步有望促进我们对遗传学在决定 mRNA 修饰中的作用的理解。