One-, two-, and three-color whole-mount in situ hybridization to Drosophila embryos

Methods : a Companion to Methods in Enzymology
G Hauptmann

Abstract

This article contains detailed protocols for the localization of mRNA transcripts within whole Drosophila embryos. The procedures are based on the use of digoxigenin-, fluorescein-, and biotin-labeled antisense RNA probes for nonradioactive detection of transcripts. The labels are visualized in situ by differently colored water-insoluble precipitates using alkaline phosphatase- or beta-galactosidase-based immunoassays. First, a basic method is described that allows detection of transcript distribution(s) of one or more genes using the same color precipitate. Second, a sequential alkaline phosphatase detection method is presented that permits the visualization of two or three independent transcript patterns in multiple colors in the same embryo. Third, a shortened two-color in situ hybridization protocol is provided that employs a combination of beta-galactosidase and alkaline phosphatase colorimetric reactions for differential detection. The two-color in situ hybridization methods work equally well in Drosophila and zebrafish embryos and may therefore also be adaptable to other species.

Citations

Jan 29, 2016·Methods : a Companion to Methods in Enzymology·Giselbert HauptmannIris Söll
Mar 3, 2004·Developmental Dynamics : an Official Publication of the American Association of Anatomists·Nathaniel DenkersTeri Jo Mauch
Jul 1, 2008·Gene Expression Patterns : GEP·Jorge Blanco, Walter J Gehring
Feb 20, 2007·Gene Expression Patterns : GEP·Hui ChenJanet D Robishaw
Sep 3, 2004·The Plant Journal : for Cell and Molecular Biology·Edouard PesquetDeborah Goffner
Aug 9, 2006·Developmental Dynamics : an Official Publication of the American Association of Anatomists·Romulo Hurtado, Takashi Mikawa
Jul 11, 2008·Archives of Insect Biochemistry and Physiology·Wenbin LiuYongping Huang
May 16, 2008·Archives of Insect Biochemistry and Physiology·Chun-che ChangWen-Chang Chang
Sep 6, 2012·The Journal of Comparative Neurology·Gilbert LauterGiselbert Hauptmann
Jun 10, 2010·The Journal of Comparative Neurology·Lars BräutigamGiselbert Hauptmann
Jul 28, 2006·Biochimica Et Biophysica Acta·Daoyong WangWei Xie
Sep 20, 2005·Gene Expression Patterns : GEP·Nicole C GriederWalter J Gehring
Apr 24, 2001·Methods : a Companion to Methods in Enzymology·T W Abraham
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Sep 12, 2003·Developmental Cell·Hervé AgaisseNorbert Perrimon
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Jan 22, 2020·Development Genes and Evolution·Gang LiuBaozhong Liu
Jan 22, 2003·Journal of Immunological Methods·Margherita Mariani, Paola Panina-Bordignon

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