A gene-driven ENU-based approach to generating an allelic series in any gene

• DOI: 10.1007/s00335-004-2379-z
• Publication Date: 2004-09-28T17:06:22Z
• Authors: Mohamed Mohideen ...; Alison Hugill; Neil Dear; Lucie Vizor; Sara Wells; Emma Horner; Shelly Fuller; Jessica Weedon; Hamish McMath; Paul Woodman; David Edwards; David Campbell; Susan Rodger; Joanne Carey; Ann Roberts; Pete Glenister; Zuzanna Lalanne; Nick Parkinson; Emma L. Coghill; Richard McKeone; Sam Cox; John Willan; Andy Greenfield; David Keays; Saffron Brady; Nigel Spurr; Ian Gray; Jackie Hunter; Steve D.M. Brown; Roger D. Cox
• Keywords: 0301 basic medicine; Alkylating Agents; Mice; 03 medical and health sciences; Ethylnitrosourea; DNA Mutational Analysis; Mutation; Animals; Alleles; Chromatography, High Pressure Liquid

N-ethyl-N-nitrosourea (ENU) introduces mutations throughout the mouse genome at relatively high efficiency. Successful high-throughput phenotype screens have been reported and alternative screens using sequence-based approaches have been proposed. For the purpose of generating an allelic series in selected genes by a sequence-based approach, we have constructed an archive of over 4000 DNA samples from individual F1 ENU-mutagenized mice paralleled by frozen sperm samples. Together with our previously reported archive, the total size now exceeds 6000 individuals. A gene-based screen of 27.4 Mbp of DNA, carried out using denaturing high-performance liquid chromatography (DHPLC), found a mutation rate of 1 in 1.01 Mbp of which 1 in 1.82 Mbp were potentially functional. Screening of whole or selected regions of genes on subsets of the archive has allowed us to identify 15 new alleles from 9 genes out of 15 tested. This is a powerful adjunct to conventional mutagenesis strategies and has the advantage of generating a variety of alleles with potentially different phenotypic outcomes that facilitate the investigation of gene function. It is now available to academic collaborators as a community resource.