In situ hybridization methods for mouse whole mounts and tissue sections with and without additional β-galactosidase staining.
In situ hybridization methods for mouse whole mounts and tissue sections with and without additional β-galactosidase staining.
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DOI：
10.1007/978-1-60327-292-6_1
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发表时间：
2014
期刊：
影响因子：
--
通讯作者：
Mishina, Yuji
中科院分区：
文献类型：
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作者：
Komatsu, Yoshihiro;Kishigami, Satoshi;Mishina, Yuji
原位杂交是一种检测形态学保存的样品中内源性mRNA序列的有效方法。我们提供原位杂交方法，该方法专门针对小鼠胚胎样品进行优化，作为整体安装和切片组织。此外，β-半乳糖苷酶（β-gal）是用于检测内源或外源基因表达的常用报告基因。结果表明，6-氯-3-吲哚基-β-D-吡喃半乳糖苷（S-gal）比5-溴-4-氯-3-吲哚基-β-D-半乳糖苷（X-gal）对β-gal活性更敏感。S-gal在β-gal活性有限的情况下是有利的，包括早期小鼠胚胎。由于S-gal的灵敏度增加以及颜色兼容性，我们成功地将使用S-gal的β-gal染色与使用DIG标记的探针的原位杂交结合在整个封片和切片中。
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.