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Mitochondrial DNA genotypes in nuclear transfer-derived cloned sheep

Nature Genetics volume 23, pages 90–93 (1999)Cite this article

Abstract

Eukaryotic cells contain two distinct genomes. One is located in the nucleus (nDNA) and is transmitted in a mendelian fashion, whereas the other is located in mitochondria (mtDNA) and is transmitted by maternal inheritance. Cloning of mammals1,2,3,4,5,6 typically has been achieved via nuclear transfer, in which a donor somatic cell is fused by electoporation with a recipient enucleated oocyte. During this whole-cell electrofusion, nDNA as well as mtDNA ought to be transferred to the oocyte7,8. Thus, the cloned progeny should harbour mtDNAs from both the donor and recipient cytoplasms, resulting in heteroplasmy. Although the confirmation of nuclear transfer has been established using somatic cell-specific nDNA markers, no similar analysis of the mtDNA genotype has been reported. We report here the origin of the mtDNA in Dolly, the first animal cloned from an established adult somatic cell line, and in nine other nuclear transfer-derived sheep generated from fetal cells. The mtDNA of each of the ten nuclear-transfer sheep was derived exclusively from recipient enucleated oocytes, with no detectable contribution from the respective somatic donor cells. Thus, although these ten sheep are authentic nuclear clones, they are in fact genetic chimaeras, containing somatic cell-derived nuclear DNA but oocyte-derived mtDNA.

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Figure 1: The sheep D-loop region.
Figure 2: PCR/RFLP analyses of the 544-bp PCR-amplified D-loop fragments, showing the maps with the predicted digestion fragment sizes, in bp, for each of the 4 indicated restriction endonucleases.

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References

  1. Cibelli, J.B. et al. Cloned transgenic calves produced from nonquiescent fetal fibroblasts. Science 280, 1256–1258 (1998).

    Article  CAS  PubMed  Google Scholar 

  2. Meng, L., Ely, J.J., Stouffer, R.L. & Wolf, D.P. Rhesus monkeys produced by nuclear transfer. Biol. Reprod. 57, 454–459 (1997).

    Article  CAS  PubMed  Google Scholar 

  3. Wakayama, T., Perry, A.C.F., Zuccotti, M., Johnson, K.R. & Yanagimachi, R. Full-term development of mice from enucleated oocytes injected with cumulus cell nuclei. Nature 394, 369–374 (1998).

    Article  CAS  PubMed  Google Scholar 

  4. Wells, D.N., Misica, P.M., Day, T.A. & Tervit, H.R. Production of cloned lambs from an established embryonic cell line: a comparison between in vivo- and in vitro-matured cytoplasts. Biol. Reprod. 57, 385–393 (1997).

    Article  CAS  PubMed  Google Scholar 

  5. Wilmut, I., Schnieke, A.E., McWhir, J., Kind, A.J. & Campbell, K.H.S. Viable offspring derived from fetal and adult mammalian cells. Nature 385, 810–813 (1997).

    Article  CAS  PubMed  Google Scholar 

  6. Schnieke, A.E. et al. Human factor IX transgenic sheep produced by transfer of nuclei from transfected fetal fibroblasts. Science 278, 2130–2133 (1997).

    Article  CAS  PubMed  Google Scholar 

  7. Gaertig, J., Kiersnowska, M. & Iftode, F. Induction of cybrid strains of Tetrahymena thermophila by electrofusion. J. Cell Sci. 89, 253–261 (1988).

    PubMed  Google Scholar 

  8. Sekirina, G.G., Bogoliubova, N.A., Antonova, N.V. & Dyban, A.P. The behaviour of mitochondria and cell integration during somatic hybridisation of sister blastomeres of the 2-cell mouse embryo. Zygote 5, 97–103 (1997).

    Article  CAS  PubMed  Google Scholar 

  9. Piko, L. & Taylor, K.D. Amounts of mitochondrial DNA and abundance of some mitochondrial gene transcripts in early mouse embryos. Dev. Biol. 123, 364–374 (1987).

    Article  CAS  PubMed  Google Scholar 

  10. Chen, X. et al. Rearranged mitochondrial genomes are present in human oocytes. Am. J. Hum. Genet. 57, 239–247 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. King, M.P. & Attardi, G. Human cells lacking mtDNA: repopulation with exogenous mitochondria by complementation. Science 246, 500–503 (1989).

    Article  CAS  PubMed  Google Scholar 

  12. Giles, R.E., Blanc, H., Cann, H.M. & Wallace, D.C. Maternal inheritance of human mitochondrial DNA. Proc. Natl Acad. Sci. USA 77, 6715–6719 (1980).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Kaneda, H. et al. Elimination of paternal mitochondrial DNA in intraspecific crosses during early mouse embryogenesis. Proc. Natl Acad. Sci. USA 92, 4542–4546 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Ankel-Simons, F. & Cummins, J.M. Misconceptions about mitochondria and mammalian fertilization: implications for theories on human evolution. Proc. Natl Acad. Sci. USA 93, 13859–13863 (1996).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Wood, N.J. & Phua, S.H. Variation in the control region sequence of the sheep mitochondrial genome. Anim. Genet. 27, 25–33 (1996).

    Article  CAS  PubMed  Google Scholar 

  16. Lightowlers, R.N., Chinnery, P.F., Turnbull, D.M. & Howell, N. Mammalian mitochondrial genetics: heredity, heteroplasmy and disease. Trends Genet. 13, 450–455 (1997).

    Article  CAS  PubMed  Google Scholar 

  17. Pinkert, C.A., Irwin, M.H., Johnson, L.W. & Moffatt, R.J. Mitochondria transfer into mouse ova by microinjection. Transgenic Res. 6, 379–383 (1997).

    Article  CAS  PubMed  Google Scholar 

  18. Shitara, H., Hayashi, J.I., Takahama, S., Kaneda, H. & Yonekawa, H. Maternal inheritance of mouse mtDNA in interspecific hybrids: segregation of the leaked paternal mtDNA followed by the prevention of subsequent paternal leakage. Genetics 148, 851–857 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  19. Cummins, J.M., Wakayama, T. & Yanagimachi, R. Fate of microinjected sperm components in the mouse oocyte and embryo. Zygote 5, 301–308 (1997).

    Article  CAS  PubMed  Google Scholar 

  20. Meirelles, F.V. & Smith, L.C. Mitochondrial genotype segregation in a mouse heteroplasmic lineage produced by embryonic karyoplast transplantation. Genetics 145, 445–451 (1997).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Meirelles, F.V. & Smith, L.C. Mitochondrial genotype segregation during preimplantation development in mouse heteroplasmic embryos. Genetics 148, 877–883 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Jenuth, J.P., Peterson, A.C., Fu, K. & Shoubridge, E.A. Random genetic drift in the female germline explains the rapid segregation of mammalian mitochondrial DNA. Nature Genet. 13, 146–151 (1996).

    Article  Google Scholar 

  23. Jenuth, J.P., Peterson, A.C. & Shoubridge, E.A. Tissue-specific selection for different mtDNA genotypes in heteroplasmic mice. Nature Genet. 16, 93–95 (1997).

    Article  CAS  PubMed  Google Scholar 

  24. Schon, E.A., Bonilla, E. & DiMauro, S. Mitochondrial DNA mutations and pathogenesis. J. Bioenerg. Biomembr. 29, 131–149 (1997).

    Article  CAS  PubMed  Google Scholar 

  25. Ciafaloni, E. et al. Widespread tissue distribution of a tRNALeu(UUR) mutation in the mitochondrial DNA of a patient with MELAS syndrome. Neurology 41, 1663–1665 (1991).

    Article  CAS  PubMed  Google Scholar 

  26. Silvestri, G. et al. Clinical features associated with the A→G transition at nucleotide 8344 of mtDNA ("MERRF" mutation). Neurology 43, 1200–1206 (1993).

    Article  CAS  PubMed  Google Scholar 

  27. Ferlin, T. et al. Segregation of the G8993 mutant mitochondrial DNA through generations and embryonic tissues in a family at risk of Leigh syndrome. J. Pediatr. 131, 447–449 (1997).

    Article  CAS  PubMed  Google Scholar 

  28. Rubenstein, D.S., Thomasma, D.C., Schon, E.A. & Zinaman, M.J. Germ-line therapy to cure mitochondrial disease: protocol and ethics of in vitro ovum nuclear transplantation. Camb. Q. Healthc. Ethics 4, 316–339 (1995).

    Article  CAS  PubMed  Google Scholar 

  29. Cohen, J. et al. Ooplasmic transfer in mature human oocytes. Mol. Hum. Reprod. 4, 269–280 (1998).

    Article  CAS  PubMed  Google Scholar 

  30. Buchanan, F.C., Littlejohn, R.P., Galloway, S.M. & Crawford, A.M. Microsatellites and associated repetitive elements in the sheep genome. Mamm. Genome 4, 258–264 (1993).

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We thank E.A. Shoubridge, M. Hirano and S.C. Silverstein for helpful discussions. This work was supported by grants from the National Institutes of Health (NS28828; NS11766; HD32062; HL52145) and the Muscular Dystrophy Association.

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Authors and Affiliations

  1. Integrated Program in Cellular, Molecular and Biophysical Studies, Columbia University College of Physicians and Surgeons, 630 West 168th Street, New York, 10032, New York, USA

    Matthew J. Evans & Cagan Gurer

  2. Department of Physiology and Cellular Biophysics, Columbia University College of Physicians and Surgeons, 630 West 168th Street, New York, 10032, New York, USA

    John D. Loike

  3. Department of Neurology, Columbia University College of Physicians and Surgeons, 630 West 168th Street, New York, 10032, New York, USA

    Eric A. Schon

  4. Genetics and Development, Columbia University College of Physicians and Surgeons, 630 West 168th Street, New York, 10032, New York, USA

    Eric A. Schon

  5. Roslin Institute, Midlothian, EH25 9PS, Roslin, Scotland

    Ian Wilmut

  6. PPL Therapeutics and Roslin Institute, Edinburgh, EH25 9PP, Roslin, UK

    Angelika E. Schnieke

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  1. Matthew J. Evans
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Correspondence to Eric A. Schon.

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Evans, M., Gurer, C., Loike, J. et al. Mitochondrial DNA genotypes in nuclear transfer-derived cloned sheep. Nat Genet 23, 90–93 (1999). https://doi.org/10.1038/12696

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