Skip to main content

Generation of Monoparental Embryos for Investigation into Genomic Imprinting

  • Protocol

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 181))

Abstract

The seminal work of McGrath and Solter (1) and independently of Surani et al. (2) in 1984 established the fundamental principle of nuclear nonequivalency; that is, chromosomes of both paternal and maternal origin are required for development to term in mammals. This was achieved through the creation of diploid reconstituted zygotes, which contained either two maternal or two paternal pronuclei. Embryos containing pronuclei exclusively of maternal or paternal origin display characteristic developmental abnormalities and fail to develop to term. This failure is partially explained by the observation that paternally and maternally derived genomes have complementary roles during embryogenesis, contributing differentially to embryonic and extraembryonic lineages (25). These reconstitutions were accomplished by nuclear transplantation and karyoplast fusion using HVJ or Sendai virus-assisted fusion (1). These experiments laid the foundation for the discovery and exploration of this unique form of non-Mendelian mammalian gene regulation whereby expression of genes and hence phenotype were dictated by the parent from whom they where inherited. This parent-of-origin phenomenon is known as genomic imprinting.

This is a preview of subscription content, log in via an institution.

Buying options

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   109.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. McGrath, J. and Solter, D. (1983) Nuclear transplantation in the mouse embryo by microsurgery and cell fusion. Science 220, 1300–1303.

    Article  PubMed  CAS  Google Scholar 

  2. Surani, M. A. H., Barton, S. C., and Norris, M. L. (1984) Development of reconstituted mouse eggs suggests imprinting of the genome during gametogenesis. Nature 308, 548–550.

    Article  PubMed  CAS  Google Scholar 

  3. Barton, S. C., Surani, M. A. H., and Norris, M. L. (1984) Role of paternal and maternal genomes in mouse development. Nature 311, 374–376.

    Article  PubMed  CAS  Google Scholar 

  4. Barton, S.C., Adams, C. A., Norris, M. L., and Surani, M. A. H. (1985) Development of gynogenetic and parthenogenetic inner cell mass and trophectoderm tissues in reconstructed blastocysts in the mouse. J. Embryol. Exp. Morphol. 90, 267–285.

    PubMed  CAS  Google Scholar 

  5. Surani, M. A. H., Barton, S. C., and Norris, M. L. (1986) Nuclear transplantation in the mouse: heritable differences between parental genomes after activation of the embryonic genome. Cell 45, 127–136.

    Article  PubMed  CAS  Google Scholar 

  6. Kono, T., Sotomara, Y., Sato, Y., and Nakahara, T. (1993) Development of androgenetic mouse embryos produced by in vitro fertilisation of enucleated oocytes. Molec. Reprod. Dev. 34, 43–46.

    Article  PubMed  CAS  Google Scholar 

  7. McGrath, J. and Solter, D. (1984) Completion of mouse embryogenesis requires both the maternal and paternal genomes. Cell 37, 170–183.

    Article  Google Scholar 

  8. O’Neill, Q. T., Rolfe, L. R., and Kaufman, M. H. (1991) Developmental potential and chromosome constitution of strontium-induced mouse parthenogenones. Molec. Reprod. Dev. 30, 214–219.

    Article  PubMed  Google Scholar 

  9. Bos-Mikich, A., Swann, K., and Whittingham, D. G. (1995) Calcium oscillations and protein synthesis inhibition synergistically activate mouse oocytes. Molec. Reprod. Dev. 41, 84–90.

    Article  PubMed  CAS  Google Scholar 

  10. Henery, C. C. and Kaufman, M. H. (1993) The incidence of aneuploidy after single pulse electroactivation of mouse oocytes. Molec. Reprod. Dev. 34, 299–307.

    Article  PubMed  CAS  Google Scholar 

  11. Moore, T. and Haigh, D. (1991) Genomic imprinting in mammalian development: a parental tug-of-war. Trends Genet. 7, 45–49.

    PubMed  CAS  Google Scholar 

  12. Sturm, K. S., Flannery, M. L., and Pedersen, R. A. (1994) Abnormal development of embryonic and extraembryonic cell lineages in parthenogenetic mouse embryos. Dev. Dynam. 210, 11–28.

    Article  Google Scholar 

  13. Spindle, A., Sturm, K. S., Flannery, M., Meneses, S. J., Wu, K., and Pedersen, R. A. (1996) Defective chorioallantoic fusion in mid-gestation lethality of parthenogenone tetraploid chimeras. Dev. Biol. 173, 447–458.

    Article  PubMed  CAS  Google Scholar 

  14. Searle, A. G. and Beechey, C. V. (1985) Non-complementation phenomena and their bearing on non-dysfunctional effects. In: Aneuploidy: Etiology and mechanisms (Dellarco, K. L., Voytele, P. E., and Hollaender, A., eds.), Plenum, New York, pp. 363–376.

    Google Scholar 

  15. Beechey, C. V. and Cattanach, B.M (1998) MRC Genetics Unit, Harwell, Oxford shire.World Wide WebSite. Genetic and Physical Imprinting Map of the Mouse (http://www.mgu.har.mrc.ac.uk/imprinting/imprinting.html).

    Google Scholar 

  16. Kaneko-Ishino, T., Kuroiwa, Y., Miyoshi, N., Kohda, T., Suzuki, R., Yokoyama, M., Viville, S., Barton, S. C., Ishino, F., and Surani, M. A. (1995) Peg1/Mest imprinted gene on chromosome 6 identified by cDNA subtraction hybridization. Nat. Genet. 11, 52–59.

    Article  PubMed  CAS  Google Scholar 

  17. Miyoshi, N., Kuriowa, Y., Kohda, T., Shitara, H., Yonekawa, H., Kaube, T., Hasegawa, H., Barton, S. C., Surani, M. A., Kaneko-Ishino, T., and Ishino, F. (1998)Identification of the Meg1/Grb10 imprinted gene on mouse proximal chromosome 11 a candidate for the Silver-Russell syndrome gene. Proc. Natl. Acad. Sci. USA 95, 1102–1107.

    Article  PubMed  CAS  Google Scholar 

  18. Kelsey, G. and Reik, W. (1998)Analysis and identification of imprinted genes.Methods 14, 211–234.

    Article  PubMed  CAS  Google Scholar 

  19. Peters, J., Wroe, S. F., Wells, C. A., Miller, H. J., Bodle, D., Beechey, C. V., Williamson, C. M., and Kelsey, G. (1999)A cluster of oppositely imprinted transcripts at the Gnas locus in the distal imprinting region of mouse chromosome 2. Proc. Natl. Acad. Sci. USA 96, 3830–3835.

    Article  PubMed  CAS  Google Scholar 

  20. Dietrich, W., Katz, H., Lincoln, S. E., Shin, H. S., Friedman, J., Pracopoli, N. C., and Lander, E. S. (1992) A genetic map of the mouse suitable for typing intraspecific crosses.Genetics 131, 423–447.

    PubMed  CAS  Google Scholar 

  21. Barton, S. C. and Surani, M. A. (1993)Manipulations of genetic constitution by nuclear transplantation.Methods Enzymol. 225, 732–744.

    Article  PubMed  CAS  Google Scholar 

  22. Eglitis, M. A. (1980)Formation of tetraploid mouse blastocysts following fusion with polyethylene glycol.J. Exp. Zool. 213, 309–313.

    Article  PubMed  CAS  Google Scholar 

  23. Kubiak, J. Z. and Tarkowski, A. K. (1985) Electrofusion of mouse blastomeres.Exp. Cell Res. 157, 561–5676.

    Article  PubMed  CAS  Google Scholar 

  24. Collas, P., Fissore, R., and Robl, J. M. (1993)Preparation of nuclear transplant embryos by electroporation.Anal. Biochem. 208, 1–9.

    Article  PubMed  CAS  Google Scholar 

  25. Tatham, B. G., Giliam, K. J., and Trounson, A. O. (1996) Electrofusion parameters for nuclear transfer predicted using isofusion contours produced with bovine embryonic cells.Molec. Reprod. Dev. 43, 306–312.

    Article  PubMed  CAS  Google Scholar 

  26. Tsuoda, Y., Kato, Y., and Shioda, Y. (1987) Electrofusion for the transplantation of mouse eggs.Gamete Res. 17, 15–20.

    Article  Google Scholar 

  27. Kono, T. and Tsunoda, Y. (1988) Nuclear transplantation of rat embryos.J. Exp. Zool. 248, 303–305.

    Article  Google Scholar 

  28. Nagy, A., Gócza, E., Diaz, E. M., Prideaux, V. R., Iványin, E., Markula, M., and Rossant, J. (1990) Embryonic stem cells alone are able to support fetal development in the mouse.Development 110, 815–821.

    PubMed  CAS  Google Scholar 

  29. McLaughlin, K. J. (1993) Production of tetraploid embryos by electrofusion.Methods Enzymol. 225, 919–930.

    Article  PubMed  CAS  Google Scholar 

  30. Lawitts, J. A. and Biggers, J. D. (1993) Culture of preimplantation embryos.Methods Enzymol. 225, 153–190.

    Article  PubMed  CAS  Google Scholar 

  31. Hogan, B., Costantini, F., and Lacy, E. (1986) Manipulating the Mouse Embryo: A Laboratory Manual.Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.

    Google Scholar 

  32. Papaioannou, V. and Johnson, R. (1993) Production of chimeras and genetically defined offspring from targeted ES cells. In: Gene Targeting: A Practical Approach (Joyner, A. L., ed.), IRL Press at Oxford University Press, Oxford, U.K., pp. 107–146.

    Google Scholar 

  33. Kaufman, M. H., Barton, S. C., Surani, M. A. H. (1977) Normal postimplantation development of mouse parthenogenetic embryos to the forelimb bud stage.Nature 265, 53–55.

    Article  PubMed  CAS  Google Scholar 

  34. Graham, C. F. (1971) Virus assisted fusion of embryonic cells.In:Karolinska Symposia on Research Methods in Reproductive Endocrinology (Diczfalusy, E., ed.), Karolinska Institute, Stockholm, Sweden, pp. 154–165.

    Google Scholar 

  35. Tarkowski, A. K., Witkowska, A., and Opas, J. (1977) Development of cytochalasin B induced tetraploid and diploid/tetraploid mosaic mouse embryos.J. Embryol. Exp. Morphol. D41, 47–64.

    Google Scholar 

  36. James, R. M., Klerkx, A. H. E. M., Keighnen, M., Flockhart, J. H., and West, J. D. (1995) Restricted distribution of tetraploid cells in mouse tetraploid ? diploid chimaeras.Dev. Biol. 167, 213–226.

    Article  PubMed  CAS  Google Scholar 

  37. Nagy, A. and Rossant, J. (1993) Production of completely ES cell derived fetuses.In: Gene Targeting: A Practical Approach (Joyner, A. L., ed.), IRL Press at Oxford University Press, Oxford, U.K., pp. 147–179.

    Google Scholar 

  38. Friedrich, G. and Soriono, P. (1991) Promoter traps in embryonic stem cells: a genetic screen to identify and mutate developmental genes in mice. Genes Dev. 5, 1513–1523.

    Article  PubMed  CAS  Google Scholar 

  39. Stewart, C. L. (1993) Production of chimeras between embryonic stem cells and embryos. Methods Enzymol. 225, 823–855.

    Article  PubMed  CAS  Google Scholar 

  40. Latham, K. E. and Solter, D. (1991) Effect of egg composition on the developmental capacity of androgenetic mouse embryos. Development 113, 561–568.

    PubMed  CAS  Google Scholar 

  41. Gossler, A. and Zachgo, J. (1993) Gene enhancer trap screens in ES cell chimeras. In: Gene Targeting: A Practical Approach (Joyner, A. L., ed.), IRL Press at Oxford University Press, Oxford, UK, pp. 181–227.

    Google Scholar 

  42. Dean, W., Bowden, L., Aitchson, A., Klose, J., Moore, T, Meneses, J. J., Reik, W., and Feil, R. (1998) Altered imprinted gene methylation and expression completely ES cell-derived mouse fetuses: association with aberrant phenotypes.Development 125, 2273–2282.

    PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2002 Humana Press Inc., Totowa, NJ

About this protocol

Cite this protocol

Dean, W.L., Kelsey, G., Reik, W. (2002). Generation of Monoparental Embryos for Investigation into Genomic Imprinting. In: Ward, A. (eds) Genomic Imprinting. Methods in Molecular Biology™, vol 181. Humana Press. https://doi.org/10.1385/1-59259-211-2:1

Download citation

  • DOI: https://doi.org/10.1385/1-59259-211-2:1

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-741-0

  • Online ISBN: 978-1-59259-211-1

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics