Production of conditional point mutant knockin mice
Production of conditional point mutant knockin mice
复制标题
DOI：
10.1002/dvg.20222
复制
发表时间：
2006-07-01
期刊：
影响因子：
1.5
通讯作者：
Homanics, Gregg E.
中科院分区：
文献类型：
--
作者：
Skvorak, Kristen;Vissel, Bryce;Homanics, Gregg E.
内源基因点突变的基因工程小鼠（即敲入小鼠）是剖析基因功能的极其有用的工具。目前用于创建敲入小鼠的可用方法是有限的，因为引入的突变普遍存在于动物从受孕到成年的所有细胞中。在本报告中，我们描述了一种创建小鼠的策略，其中点突变等位基因以有条件的方式（例如，以组织特异性和/或时间限制的模式）取代野生型等位基因。作为概念证明，我们创造了有条件地携带点突变的γ-氨基丁酸受体亚基的小鼠。在没有 Cre 重组酶的情况下，工程等位基因仅产生野生型产物，没有突变体表达的证据。相反，在Cremediad重组之后，仅产生点突变产物。通过将 Cre 表达限制在出生后动物的神经元亚群中，我们证明了点突变敲入的组织特异性调节。该策略可用于需要用点突变体精确条件替换内源野生型基因的各种研究。
Genetically engineered mice with point mutations in endogenous genes (i.e., knockin mice) are extremely useful tools for dissecting gene function. Currently available methodologies for creating knockin mice are limited in that the introduced mutation is globally present in all cells of the animal from conception through adulthood. In this report, we describe a strategy for creating mice in which a point mutant allele replaces the wild type allele in a conditional manner, e.g., in a tissue-specific and/or temporally restricted pattern. As proof of concept, we created mice that conditionally harbor a point mutated gamma-aminobutyric acid receptor subunit. In the absence of Cre recombinase, the engineered allele produces only wild type product with no evidence of expression of the mutant. In contrast, following Cremediated recombination, only the point mutant product is produced. By restricting Cre expression to subpopulations of neurons of postnatal animals, we demonstrate tissuespecific regulation of the point mutant knockin. This strategy will be useful for a wide variety of studies that require precise conditional replacement of an endogenous wild type gene with a point mutant.