Whole-genome high-fidelity sequencing: A novel approach to detecting and characterization of mutagenicity in vivo

Direct DNA sequencing can be used for characterizing mutagenicity in simple and complex biological models. Recently we described a method of whole-genome sequencing for detecting mutations in simple models of cultured bacteria, mammalian cells, and nematode. In the current proof-of-concept study, we expand and improve our method for evaluating a more complex mammalian biological model in outbred mice. We detail the method by applying it to a small set of animals treated with a mutagen with known mutagenicity profiles, N-ethyl-N-nitrosourea (ENU), for consistency with the known data. Whole-genome high-fidelity sequencing (HiFi Sequencing) showed frequencies and spectra of background mutations in tissues of untreated mice that were consistent with normal ageing and characterized by spontaneous or enzymatic deamination of 5-methylcytosine. In mice treated with a single 40 mg/kg dose of ENU, the frequency of mutations in the genomic DNA of solid tissues increased up to 7-fold, with the greatest increase observed in the spleen and the smallest increase in the liver. The most common mutations detected in ENU-treated mice were T > A transitions and T > C transversions, consistent with the types of mutations caused by alkylating agents. The data suggest that HiFi Sequencing may be useful for characterizing mutagenicity of novel compounds in various biological models. •Mutations are involved in cancer onset and progression.•High-Fidelity Sequencing detects mutations induced by prototypical mutagens.•HiFi Sequencing may be useful for identification of potential in vivo carcinogens.