Mouse mutants from chemically mutagenized embryonic stem cells (original) (raw)

Nature Genetics volume 24, pages 318–321 (2000)Cite this article

Abstract

The drive to characterize functions of human genes on a global scale has stimulated interest in large-scale generation of mouse mutants. Conventional germ-cell mutagenesis with _N_-ethyl-_N_-nitrosourea (ENU) is compromised by an inability to monitor mutation efficiency, strain1 and interlocus2 variation in mutation induction, and extensive husbandry requirements. To overcome these obstacles and develop new methods for generating mouse mutants, we devised protocols to generate germline chimaeric mice from embryonic stem (ES) cells heavily mutagenized with ethylmethanesulphonate (EMS). Germline chimaeras were derived from cultures that underwent a mutation rate of up to 1 in 1,200 at the Hprt locus (encoding hypoxanthine guanine phosphoribosyl transferase). The spectrum of mutations induced by EMS and the frameshift mutagen ICR191 was consistent with that observed in other mammalian cells. Chimaeras derived from ES cells treated with EMS transmitted mutations affecting several processes, including limb development, hair growth, hearing and gametogenesis. This technology affords several advantages over traditional mutagenesis, including the ability to conduct shortened breeding schemes and to screen for mutant phenotypes directly in ES cells or their differentiated derivatives.

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References

  1. Bode, V.C. Ethylnitrosourea mutagenesis and the isolation of mutant alleles for specific genes located in the T region of mouse chromosome 17. Genetics 108, 457–470 ( 1984).
    CAS PubMed PubMed Central Google Scholar
  2. Rinchik, E.M. & Carpenter, D.A. N-Ethyl-N-nitrosourea mutagenesis of a 6- to 11-cM subregion of the Fah-Hbb interval of mouse chromosome 7. Completed testing of 4557 gametes and deletion mapping and complementation analysis of 31 mutations. Genetics 152, 373–383 (1999).
    CAS PubMed PubMed Central Google Scholar
  3. Sega, G.A. A review of the genetic effects of ethyl methanesulfonate. Mutat. Res. 134, 113–142 ( 1984).
    Article CAS PubMed Google Scholar
  4. Klungland, A., Laake, K., Hoff, E. & Seeberg, E. Spectrum of mutations induced by methyl and ethyl methanesulfonate at the hprt locus of normal and tag expressing Chinese hamster fibroblasts. Carcinogenesis 16, 1281–1285 (1995).
    Article CAS PubMed Google Scholar
  5. Hsie, A.W., Brimer, P.A., Mitchell, T.J. & Gosslee, D.G. The dose-response relationship for ethyl methanesulfonate-induced mutations at the hypoxanthine-guanine phosphoribosyl transferase locus in Chinese hamster ovary cells. Somat. Cell Genet. 1, 247– 261 (1975).
    Article CAS PubMed Google Scholar
  6. Taft, S.A., Liber, H.L. & Skopek, T.R. Mutational spectrum of ICR-191 at the hprt locus in human lymphoblastoid cells. Environ. Mol. Mutagen 23, 96–100 (1994).
    Article CAS PubMed Google Scholar
  7. Chen, Y. et al. Genotype-based screen for ENU-induced mutations in mouse embryonic stem cells. Nature Genet. 24, 314– 317 (2000).
    Article CAS PubMed Google Scholar
  8. You, Y. et al. Chromosomal deletion complexes in mice by radiation of embryonic stem cells. Nature Genet. 15, 285– 288 (1997).
    Article CAS PubMed Google Scholar
  9. Ramirez Solis, R., Liu, P. & Bradley, A. Chromosome engineering in mice. Nature 378, 720–724 (1995).
    Article CAS PubMed Google Scholar
  10. Zheng, Q.Y., Johnson, K.R. & Erway, L.C. Assessment of hearing in 80 inbred strains of mice by ABR threshold analyses. Hear. Res. 130, 94–107 (1999).
    Article CAS PubMed PubMed Central Google Scholar
  11. John, S.W. et al. Essential iris atrophy, pigment dispersion, and glaucoma in DBA/2J mice. Invest. Ophthalmol. Vis. Sci. 39, 951–962 (1998).
    CAS PubMed Google Scholar
  12. Smith, R.S., Roderick, T.H. & Sundberg, J.P. Microphthalmia and associated abnormalities in inbred black mice. Lab. Anim. Sci. 44, 551– 560 (1994).
    CAS PubMed Google Scholar
  13. Kessel, M. & Gruss, P. Homeotic transformations of murine vertebrae and concomitant alteration of Hox codes induced by retinoic acid . Cell 67, 89–104 (1991).
    Article CAS PubMed Google Scholar

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Acknowledgements

We thank T. Magnuson for CT129 ES cells; T. Gridley for CJ7 ES cells; A. Planchart for input on design of culture experiments; J. Sztein for assistance with sperm examination; C. O'Neill for animal care; and T. O'Brien, G. Cox, T. Magnuson and M.A. Handel for critical evaluation of the manuscript. This work was supported by NIH grant GM45415 to J.C.S. and a Cancer Center Grant CA34196 to The Jackson Laboratory V.L.B. was supported by an NIH training grant (HD07065) and fellowship NIH (HD08441). S.W.M.J. is an Assistant Investigator of The Howard Hughes Medical Institute. Hearing tests were performed under a contract (DC62108) to K. Johnson from the National Institute on Deafness and Other Communicative Disorders.

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Authors and Affiliations

  1. The Jackson Laboratory, Bar Harbor, Maine, USA
    Robert J. Munroe, Rebecca A. Bergstrom, Qing Y Zheng, Brian Libby, Richard Smith, Simon W.M. John, Kerry J. Schimenti, Victoria L. Browning & John C. Schimenti
  2. The Howard Hughes Medical Institute, Bar Harbor, Maine, USA
    Richard Smith & Simon W.M. John

Authors

  1. Robert J. Munroe
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  2. Rebecca A. Bergstrom
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  3. Qing Y Zheng
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  4. Brian Libby
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  5. Richard Smith
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  6. Simon W.M. John
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  7. Kerry J. Schimenti
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  8. Victoria L. Browning
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  9. John C. Schimenti
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Corresponding author

Correspondence toJohn C. Schimenti.

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Munroe, R., Bergstrom, R., Y Zheng, Q. et al. Mouse mutants from chemically mutagenized embryonic stem cells.Nat Genet 24, 318–321 (2000). https://doi.org/10.1038/73563

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