Sakane Yuto, Suzuki Ken-Ich T, Yamamoto Takashi
Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, 1-3-1 Kagamiyama, Higashi-Hiroshima, Hiroshima, 739-8526, Japan.
Methods Mol Biol. 2017;1630:189-203. doi: 10.1007/978-1-4939-7128-2_16.
Xenopus tropicalis is a versatile model organism for studying basic biology such as developmental biology and cell biology, and for biomedical research on human diseases. Current genome editing techniques enable researchers to easily perform gene targeting in various animals. Among them, gene knockout using the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-CRISPR-associated (Cas) (CRISPR-Cas) system has recently become an indispensable strategy for loss-of-function analysis in vivo. Because of its ease of use, time, and cost efficiencies, CRISPR-Cas has also been applied to X. tropicalis where the gene disruption is highly efficient. In this chapter, we introduce a simple CRISPR-Cas system protocol for gene disruption in X. tropicalis. Based on our protocol, researchers can generate knock-out phenotypes within the shortest of timeframes, a week, and analyze genes of interest in founder generation.
热带爪蟾是一种用于研究基础生物学（如发育生物学和细胞生物学）以及人类疾病生物医学研究的通用模式生物。当前的基因组编辑技术使研究人员能够轻松地在各种动物中进行基因靶向。其中，使用成簇规律间隔短回文重复序列（CRISPR）-CRISPR相关蛋白（Cas）（CRISPR-Cas）系统进行基因敲除最近已成为体内功能丧失分析中不可或缺的策略。由于其使用方便、省时且成本效益高，CRISPR-Cas也已应用于热带爪蟾，在那里基因破坏效率很高。在本章中，我们介绍一种用于热带爪蟾基因破坏的简单CRISPR-Cas系统方案。根据我们的方案，研究人员可以在最短的时间内，即一周内产生敲除表型，并在奠基者一代中分析感兴趣的基因。
