Whole Mount In Situ Hybridization of Mid-Gestation Mouse Embryos

Q1 Agricultural and Biological Sciences

Current protocols in mouse biology Pub Date : 2020-06-15 DOI:10.1002/cpmo.77

Annemiek Beverdam

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引用次数: 1

Abstract

In situ hybridization is a powerful technique that allows the visualization of specific RNA species in biological samples in exquisite detail. It has been particularly well explored in the field of developmental genetics. The spatial and temporal patterns of RNA expression provide us with critical information on likely gene function during embryonic development, and often inform the decision on whether to attempt further gene manipulation approaches. Furthermore, once a mouse strain with altered gene function has been created, in situ hybridization is a critical tool for revealing how the development of embryos with the mutation differs from that of wild-type embryos, and thus infer the function of the altered gene. Here, a well-tested protocol used to visualize RNA expression in whole-mount mid-gestation mouse embryos ranging from 8.5 to 14.5 days post-coitum (dpc) is described. © 2020 Wiley Periodicals LLC.

Basic Protocol 1: RNA probe synthesis

Alternate Protocol: Preparation of DNA template by PCR

Basic Protocol 2: Embryo dissection

Basic Protocol 3: Whole mount in situ hybridization

Support Protocol: Generation of embryo powder

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中期妊娠小鼠胚胎全坐骨原位杂交

原位杂交是一种强大的技术，它可以使生物样品中特定RNA物种的精细细节可视化。它在发育遗传学领域已经得到了很好的探索。RNA表达的时空模式为我们提供了胚胎发育过程中可能的基因功能的关键信息，并经常告知是否尝试进一步的基因操作方法的决定。此外，一旦创建了具有改变基因功能的小鼠品系，原位杂交是揭示具有突变的胚胎与野生型胚胎的发育差异的关键工具，从而推断改变基因的功能。本文描述了一种经过充分测试的方案，用于在性交后8.5至14.5天(dpc)的全载妊娠中期小鼠胚胎中可视化RNA表达。©2020 Wiley期刊有限责任公司基本方案1:RNA探针合成备用方案:通过pcr制备DNA模板基本方案2:胚胎解剖基本方案3:全mount原位杂交支持方案:胚胎粉的产生

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Current protocols in mouse biology Agricultural and Biological Sciences-Animal Science and Zoology

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期刊介绍： Sound and reproducible laboratory methods are the foundation of scientific discovery. Yet, all too often, nuances that are critical for an experiment''s success are not captured in the primary literature but exist only as part of a lab''s oral tradition. The aim of Current Protocols in Mouse Biology is to provide the clearest, most detailed and reliable step-by-step instructions for protocols involving the use of mice in biomedical research. Written by experts in the field and extensively edited to our exacting standards, the protocols include all of the information necessary to complete an experiment in the laboratory—introduction, materials lists with supplier information, detailed step-by-step procedures with helpful annotations, recipes for reagents and solutions, illustrative figures and information-packed tables. Each article also provides invaluable discussions of background information, applications of the methods, important assumptions, key parameters, time considerations, and tips to help avoid common pitfalls and troubleshoot experiments. Furthermore, Current Protocols in Mouse Biology content is thoughtfully organized by topic for optimal usage and to maximize contextual knowledge. Quarterly issues allow Current Protocols to constantly evolve to keep pace with the newest discoveries and developments. Current Protocols in Mouse Biology brings together resources in mouse biology and genetics and provides a mouse protocol resource that covers all aspects of mouse biology. Current Protocols in Mouse Biology also permits optimization of mouse model usage, which is significantly impacted by both cost and ethical constraints. Optimal and standardized mouse protocols ultimately reduce experimental variability and reduce the number of animals used in mouse experiments.