Abstract
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.
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Acknowledgments
This work was supported by a European Union FP5 Grant INFRAQTL contract number QLRI-CT-2000-00233 (sperm freezing, DNA preparation, and selected gene screens), EUMORPHIA, The Medical Research Council, GlaxoSmithKline (GSK–Harwell portion of the archive), and Etiologics Ltd (new portion of archive). DK was supported by the Christopher Welch Trust. S. Brady was supported by the Wellcome Trust.
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Quwailid, M.M., Hugill, A., Dear, N. et al. A gene-driven ENU-based approach to generating an allelic series in any gene. Mamm Genome 15, 585–591 (2004). https://doi.org/10.1007/s00335-004-2379-z
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DOI: https://doi.org/10.1007/s00335-004-2379-z