Lintott Lauri G, Nutter Lauryl M J
The Centre for Phenogenomics, Toronto, ON, Canada.
The Hospital for Sick Children, Toronto, ON, Canada.
Methods Mol Biol. 2023;2631:53-101. doi: 10.1007/978-1-0716-2990-1_3.
Genetically engineered mice are used as avatars to understand mammalian gene function and develop therapies for human disease. During genetic modification, unintended changes can occur, and these changes may result in misassigned gene-phenotype relationships leading to incorrect or incomplete experimental interpretations. The types of unintended changes that may occur depend on the allele type being made and the genetic engineering approach used. Here we broadly categorize allele types as deletions, insertions, base changes, and transgenes derived from engineered embryonic stem (ES) cells or edited mouse embryos. However, the methods we describe can be adapted to other allele types and engineering strategies. We describe the sources and consequ ences of common unintended changes and best practices for detecting both intended and unintended changes by screening and genetic and molecular quality control (QC) of chimeras, founders, and their progeny. Employing these practices, along with careful allele design and good colony management, will increase the chance that investigations using genetically engineered mice will produce high-quality reproducible results, to enable a robust understanding of gene function, human disease etiology, and therapeutic development.
基因工程小鼠被用作替身，以了解哺乳动物基因功能并开发人类疾病的治疗方法。在基因改造过程中，可能会出现意外变化，这些变化可能导致基因-表型关系错误分配，从而导致实验解释不正确或不完整。可能出现的意外变化类型取决于所制造的等位基因类型和所使用的基因工程方法。在这里，我们将等位基因类型大致分为缺失、插入、碱基变化以及源自工程化胚胎干细胞（ES）或编辑过的小鼠胚胎的转基因。然而，我们描述的方法可适用于其他等位基因类型和工程策略。我们描述了常见意外变化的来源和后果，以及通过对嵌合体、奠基者及其后代进行筛选以及遗传和分子质量控制（QC）来检测预期和意外变化的最佳实践。采用这些做法，再加上精心的等位基因设计和良好的群体管理，将增加使用基因工程小鼠进行的研究产生高质量可重复结果的机会，从而能够深入了解基因功能、人类疾病病因和治疗开发。
