title: Current strategies for mutation detection in phenotype-driven screens utilising next generation sequencing.
abstract: Mutagenesis-based screens in mice are a powerful discovery platform to identify novel genes or gene functions associated with disease phenotypes. An N-ethyl-N-nitrosourea (ENU) mutagenesis screen induces single nucleotide variants randomly in the mouse genome. Subsequent phenotyping of mutant and wildtype mice enables the identification of mutated pathways resulting in phenotypes associated with a particular ENU lesion. This unbiased approach to gene discovery conducts the phenotyping with no prior knowledge of the functional mutations. Before the advent of affordable next generation sequencing (NGS), ENU variant identification was a limiting step in gene characterization, akin to 'finding a needle in a haystack'. The emergence of a reliable reference genome alongside advances in NGS has propelled ENU mutation discovery from an arduous, time-consuming exercise to an effective and rapid form of mutation discovery. This has permitted large mouse facilities worldwide to use ENU for novel mutation discovery in a high-throughput manner, helping to accelerate basic science at the mechanistic level. Here, we describe three different strategies used to identify ENU variants from NGS data and some of the subsequent steps for mutation characterisation.
authors.affiliation: Medical Research Council Harwell (Mammalian Genetics Unit and Mary Lyon Centre), Harwell Campus, Oxfordshire, OX11 0RD, UK. m.simon@har.mrc.ac.uk.
authors.forename: Michelle M
authors.initials: MM
authors.lastname: Simon
authors.affiliation: Center for the Genetics of Host Defense, University of Texas Southwestern Medical Center, Dallas, TX, 75390, USA.
authors.forename: Eva Marie Y
authors.initials: EM
authors.lastname: Moresco
authors.affiliation: Nuffield Department of Medicine and Wellcome Trust Centre for Human Genetics, Oxford University, Oxford, UK.|MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, Oxford, UK.
authors.forename: Katherine R
authors.initials: KR
authors.lastname: Bull
authors.affiliation: Medical Research Council Harwell (Mammalian Genetics Unit and Mary Lyon Centre), Harwell Campus, Oxfordshire, OX11 0RD, UK.
authors.forename: Saumya
authors.initials: S
authors.lastname: Kumar
authors.affiliation: Medical Research Council Harwell (Mammalian Genetics Unit and Mary Lyon Centre), Harwell Campus, Oxfordshire, OX11 0RD, UK.
authors.forename: Ann-Marie
authors.initials: AM
authors.lastname: Mallon
authors.affiliation: Center for the Genetics of Host Defense, University of Texas Southwestern Medical Center, Dallas, TX, 75390, USA.
authors.forename: Bruce
authors.initials: B
authors.lastname: Beutler
authors.affiliation: Medical Research Council Harwell (Mammalian Genetics Unit and Mary Lyon Centre), Harwell Campus, Oxfordshire, OX11 0RD, UK.
authors.forename: Paul K
authors.initials: PK
authors.lastname: Potter
mesh_terms: D000818:Animals; D004195:Disease Models, Animal; D005038:Ethylnitrosourea / Q000633:toxicity; D016678:Genome / Q000187:drug effects*; D059014:High-Throughput Nucleotide Sequencing; D051379:Mice; D016296:Mutagenesis / Q000187:drug effects / Q000235:genetics; D009154:Mutation / Q000187:drug effects* / Q000235:genetics
chemical_list: D005038:Ethylnitrosourea