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Methods Mol Biol. 2014;1092:1-15. doi: 10.1007/978-1-60327-292-6_1.

In situ hybridization methods for mouse whole mounts and tissue sections with and without additional β-galactosidase staining.

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School of Biology-Oriented Science and Technology, Kinki University, Kinki, Japan.


In situ hybridization is a powerful method for detecting endogenous mRNA sequences in morphologically preserved samples. We provide in situ hybridization methods, which are specifically optimized for mouse embryonic samples as whole mounts and section tissues. Additionally, β-Galactosidase (β-gal) is a popular reporter for detecting the expression of endogenous or exogenous genes. We reveal that 6-chloro-3-indoxyl-β-D-galactopyranoside (S-gal) is a more sensitive substrate for β-gal activity than 5-bromo-4-chloro-3-indolyl-β-D-galactoside (X-gal). S-gal is advantageous where β-gal activity is limited including early stage mouse embryos. As a result of the increased sensitivity as well as the color compatibility of S-gal, we successfully combined β-gal staining using S-gal with in situ hybridization using DIG-labeled probes in both whole mounts and sections.

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