Nanopore sequencing of RNA modifications

2020 Fiscal Year Final Research Report

• Project/Area Number: 19K22246
• Research Category: Grant-in-Aid for Challenging Research (Exploratory)
• Allocation Type: Multi-year Fund
• Review Section: Medium-sized Section 37:Biomolecular chemistry and related fields
• Research Institution: The University of Tokyo
• Principal Investigator: SUZUKI Tsutomu 東京大学, 大学院工学系研究科(工学部), 教授 (20292782)
• Project Period (FY): 2019-06-28 – 2021-03-31
• Keywords: RNA修飾 / tRNA / エピトランスクリプトミクス / ナノポアシーケンス

Outline of Final Research Achievements

For expanding world of epitranscriptomics, it is necessary to establish an innovative technology to profile and analyze cellular tRNAs with their modification status. Nanopore-based sequencing is a unique method capable of directly analyzing RNA molecules without cDNA conversion. We aim to establish a practical method for classifying cellular tRNAs with their modification status using the ONT’s nanopore sequencer. We isolated all 44 species of E. coli tRNAs and nanopore-sequenced each of them individually. All tRNA datasets were used for deep learning using a convolutional neural network (CNN). After examining various learning models and increasing the number of epochs, we succeeded in raising the classification accuracy to a practical level (>98% on average). In addition, we successfully discriminated tRNAs with or without single modification with high accuracy, demonstrating that the nanopore clearly recognizes the change in ion current signal caused by a single RNA modification.

Free Research Field

分子生物学

Academic Significance and Societal Importance of the Research Achievements

本研究成果は、エピトランスクリプトミクス研究の基盤技術を提供し、RNA修飾が担う機能と、普遍的な生命現象との関わりを明らかにすることで、学術的に貢献するだけでな、将来的なエピトランスクリプトミクス創薬や食料問題など、様々な経済的価値の創出に大きく貢献することが期待される。特に本技術は、がんや生活習慣病などの疾患で変動するRNA修飾を、微量な試料を用いて、迅速にかつ正確に解析することが可能になるため、有用なバイオマーカーが存在しない疾患領域においても、新しい診断法を提供できる可能性があると考えられる。