Abstract
Whole-mount in situ hybridization (WISH) is widely used to visualize the site and dynamics of gene expression during embryonic development. Various methods of probe labeling and hybridization detection are available nowadays. Meanwhile the technique was adapted to be used on many different species and has evolved from a manual to a larger scale and automated procedure. Standardized automated protocols improve the chance to compare different experimental settings reliably. The high resolution of this method is ideally suited for examination of manipulated (e.g., cloned) embryos often displaying subtle changes only. Embedding and sectioning of in situ hybridized specimen further enhance the detailed examination of their gene expression and morphology.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Tautz D, Pfeifle C (1989) A non-radioactive in situ hybridization method for the localization of specific RNAs in Drosophila embryos reveals translational control of the segmentation gene hunchback. Chromosoma 98(2):81–85
Hue I, Renard JP, Viebahn C (2001) Brachyury is expressed in gastrulating bovine embryos well ahead of implantation. Dev Genes Evol 211(3):157–159
Idkowiak J, Weisheit G, Plitzner J, Viebahn C (2004) Hypoblast controls mesoderm generation and axial patterning in the gastrulating rabbit embryo. Dev Genes Evol 214(12):591–605
Idkowiak J, Weisheit G, Viebahn C (2004) Polarity in the rabbit embryo. Semin Cell Dev Biol 15(5):607–617
Guillomot M, Turbe A, Hue I, Renard JP (2004) Staging of ovine embryos and expression of the T-box genes Brachyury and Eomesodermin around gastrulation. Reproduction 127(4):491–501
Hassoun R, Schwartz P, Feistel K, Blum M, Viebahn C (2009) Axial differentiation and early gastrulation stages of the pig embryo. Differentiation 78(5):301–311
Herrmann BG (1991) Expression pattern of the Brachyury gene in whole-mount TWis/TWis mutant embryos. Development 113(3):913–917
Jouneau A et al (2006) Developmental abnormalities of NT mouse embryos appear early after implantation. Development 133(8):1597–1607
Rielland M, Brochard V, Lacroix MC, Renard JP, Jouneau A (2009) Early alteration of the self-renewal/differentiation threshold in trophoblast stem cells derived from mouse embryos after nuclear transfer. (Translated from eng). Dev Biol 334(2):325–334
Degrelle SA et al (2012) Uncoupled embryonic and extra-embryonic tissues compromise blastocyst development after somatic cell nuclear transfer. PLoS One 7(6):e38309
Nakamura T, Hamada H (2013) Fluorescent 2 color Whole mount in situ hybridization for a mouse embryo. Protocol Exchange doi:10.1038/protex.2013.002
Acloque H, Wilkinson DG, Nieto MA (2008) In situ hybridization analysis of chick embryos in whole-mount and tissue sections. Methods Cell Biol 87:169–185
Brend T, Holley SA (2009) Zebrafish whole mount high-resolution double fluorescent in situ hybridization. J Vis Exp doi: 10.3791/1229
Püschel B, Blum M, Viebahn C (2010) Whole-mount in situ hybridization of early rabbit embryos. Cold Spring Harb Protoc (1):pdb prot5355
Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY
Weisheit G, Mertz D, Schilling K, Viebahn C (2002) An efficient in situ hybridization protocol for multiple tissue sections and probes on miniaturized slides. Dev Genes Evol 212(8):403–406
Hue I, Degrelle SA, Viebahn C (2013) Analysis of molecular markers for staging peri-gastrulating bovine embryos. Methods Mol Biol 1074:125–135
Püschel B, Viebahn C (2010) Rabbit mating and embryo isolation. Cold Spring Harb Protoc (1):pdb prot5350
Degrelle SA et al (2005) Molecular evidence for a critical period in mural trophoblast development in bovine blastocysts. Dev Biol 288(2):448–460
Hogan B, Beddington R, Costantini F, Lacy E (1994) Manipulating the mouse embryo: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY
Shimizu H, Uchibe K, Asahara H (2009) Large-scale whole mount in situ hybridization of mouse embryos. Methods Mol Biol 577:167–179
Hopf C, Viebahn C, Püschel B (2011) BMP signals and the transcriptional repressor BLIMP1 during germline segregation in the mammalian embryo. Dev Genes Evol 221(4):209–223
Acknowledgments
The authors’ research has been supported in part by grants as specified in our publications and by INRA and the University of Göttingen. We gratefully appreciate our colleagues for inspiring and helpful input (in Göttingen: Christoph Viebahn; in Jouy: Evelyne Campion) and for their excellent and dedicated help generating nice WISH data (in Göttingen: Kirsten Falk-Stietenroth, Heike Faust; and Irmgard Weiss, in Jouy: Vincent Brochard).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Püschel, B., Jouneau, A. (2015). Whole-Mount In Situ Hybridization to Assess Advancement of Development and Embryo Morphology. In: Beaujean, N., Jammes, H., Jouneau, A. (eds) Nuclear Reprogramming. Methods in Molecular Biology, vol 1222. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1594-1_19
Download citation
DOI: https://doi.org/10.1007/978-1-4939-1594-1_19
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-1593-4
Online ISBN: 978-1-4939-1594-1
eBook Packages: Springer Protocols