A gene-driven ENU-based approach to generating an allelic series in any gene (original) (raw)

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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Authors and Affiliations

  1. MRC Mammalian Genetics Unit, Medical Research Council, Harwell, Oxfordshire, OX11 0RD, UK
    Mohamed Mohideen Quwailid, 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, Steve D.M. Brown & Roger D. Cox
  2. Psychiatric Genetics Laboratory, The Wellcome Trust Centre for Human Genetics, Oxford University, Oxford OX3, 7BN, UK
    David Keays & Saffron Brady
  3. Discovery and Pipeline Genetics, GIaxoSmithKline Pharmaceuticals, Research Triangle Park, North Carolina, 27709, USA
    Nigel Spurr & Ian Gray
  4. GlaxoSmithKline Pharmaceuticals, New Frontiers Science Park, Harlow, CM19, 5AW, UK
    Jackie Hunter

Authors

  1. Mohamed Mohideen Quwailid
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  2. Alison Hugill
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  3. Neil Dear
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  4. Lucie Vizor
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  5. Sara Wells
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  6. Emma Horner
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  7. Shelly Fuller
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  8. Jessica Weedon
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  9. Hamish McMath
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  10. Paul Woodman
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  11. David Edwards
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  12. David Campbell
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  13. Susan Rodger
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  14. Joanne Carey
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  15. Ann Roberts
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  16. Pete Glenister
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  17. Zuzanna Lalanne
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  18. Nick Parkinson
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  19. Emma L. Coghill
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  20. Richard McKeone
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  21. Sam Cox
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  22. John Willan
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  23. Andy Greenfield
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  24. David Keays
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  25. Saffron Brady
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  26. Nigel Spurr
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  27. Ian Gray
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  28. Jackie Hunter
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  29. Steve D.M. Brown
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  30. Roger D. Cox
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Corresponding author

Correspondence toRoger D. Cox.

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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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