Development of deep-sequencing approach for genome damage repair

2016 Fiscal Year Final Research Report

• Project/Area Number: 26640117
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Genome biology
• Research Institution: The University of Tokyo (2015-2016); National Institute of Genetics (2014)
• Principal Investigator: Iida Tetsushi 東京大学, 分子細胞生物学研究所, 助教 (60391851)
• Project Period (FY): 2014-04-01 – 2017-03-31
• Keywords: ゲノム維持修復 / リボヌクレオチド / rNMP / 損傷 / 定量 / 次世代シークエンサー

Outline of Final Research Achievements

Ribonucleotides misincorporated in the genome are an inevitable source of endogenous DNA damage, and also serve as signals for genome repair. However, damage repair has been previously evaluated indirectly, and the balance between incorporation and repair of ribonucleotides has not been elucidated. In this project, we developed a competitive sequencing method, RiSQ-seq, which enables absolute quantification of misincorporated ribonucleotides throughout the genome at single-base resolution. RiSQ-seq analysis revealed that ribonucleotides were incorporated non-uniformly in the genome. Direct comparison of ribonucleotide levels in wild-type and repair-deficient mutant cells enabled evaluation of ribonucleotide excision repair (RER) activity at base resolution. Profiling of RER efficiency revealed that the distinct preferences of ribonucleotide incorporation and RER result in hotspots of insertion and deletion mutation. RiSQ-seq can be used to evaluate the risk of mutations.

Free Research Field

分子生物学