Part of the Methods in Molecular Biology™ book series (MIMB, volume 137)
Whole Mount In Situ Hybridization to Study Gene Expression During Mouse Development
Whole mount in situ hybridization (WMISH) is a method for detecting specific messenger ribonucleic acids (mRNAs) at their site of expression within an embryo or intact tissue fragment. Like conventional in situ hybridization, WMISH depends upon the availability of a complementary nucleic acid probe which can be labeled, annealed to the mRNA fixed within the tissue, and subsequently detected. Whereas the conventional approach uses radioactive probes hybridized to tissue sections, WMISH refers to hybridization to intact tissue or embryos using nonradioactive probes that are detected using standard immunohistochemical methods. For developmental biologists, WMISH offers the advantage of visualizing the domain of expression of a gene within the context of the entire embryo. In addition, WMISH can be used to study more than one gene in the same embryo, allowing spatial and temporal overlaps in expression to be clearly discerned (Fig 1). Similar to conventional in situ hybridization, one can determine the precise cellular distribution of expression by embedding and sectioning embryos following WMISH. For these reasons WMISH has become an essential and standard tool for studying gene expression during embryonic development (e.g., see 1).
KeywordsHybridization Buffer Alkaline Phosphatase Substrate Paraformaldehyde Fixative Fine Science Tool Silanated Glass Slide
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- 3.Conlon, R. A. and Herrmann, B. G. (1993) Detection of messenger RNA by in situ hybridization to postimplantation embryo whole mounts, in Methods in Enzymology, vol. 225, Guide to Techniques in Mouse Development (Wassarman, P. M. and DePamphilis, M. L., eds.), Academic, San Diego,CA, pp. 373–383.CrossRefGoogle Scholar
- 4.Hogan, B., Beddington, R., Costantini, F., and Lacy, E. (1994) Manipulating the Mouse Embryo. A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory, Plainview,NY.Google Scholar
- 8.Conlon, R. A. (1997) Whole mount in situ hybridization to embryos and embryonic tissues, in Methods in Molecular Biology, vol. 63, Recombinant Protein Protocols: Detection and Isolation(Tuan, R. S., ed.), Humana Press, Totowa, NJ, pp. 257–262.Google Scholar
- 9.Hsu, C. W. and Tuan, R. S. (1997) Whole-mount in situ hybridization for developing chick embryos using digoxygenin-labeled RNA probes, in Methods in Molecular Biology, vol. 63, Recombinant Protein Protocols: Detection and Isolation (Tuan, R. S., ed.), Humana Press, Totowa, NJ, pp. 263–270.Google Scholar
- 12.Cockroft, D. L. (1990) Dissection and culture of postimplantation embryos, in PostimplantationMammalian Embryos-A Practical Approach, (Copp, A. J. and Cockroft, D. L., eds.), IRL, Oxford, UK, pp. 15–39.Google Scholar
- 13.Sturm, K. and Tam, P. P. L. (1993) Isolation and culture of whole postimplantation embryos and germ layer derivatives, in Methods in Enzymology, vol. 225, Guide to Techniques in Mouse Development (Wassarman, P. M. and DePamphilis, M. L., eds.), Academic, San Diego, CA, pp. 164–190.CrossRefGoogle Scholar
- 14.Martins-Green, M. M. (1990) Transmission electron microscopy and immunolabelling for light and electron microscopy, in Postimplantation Mammalian Embryos-A Practical Approach, (Copp, A. J. and Cockroft, D. L.,eds.), IRL, Oxford, UK, pp. 127–154.Google Scholar
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