Loss of function mouse models comprise knock-out mice, where a gene is deleted in the germline, and conditional knock-out mice with somatic deletion of a floxed allele in defined tissues. Both types of mice are used for comprehensive studies of gene functions in vivo. Here, we describe a simple method for simultaneous generation of mice with conditional or knock-out alleles for the transcription factor fra-2 (Fos-related antigen 2) using a single embryonic stem (ES) cell clone. ES cells with a floxed fra-2 allele were transiently transfected with a Cre-recombinase expression plasmid and plated at low density. Most of the resulting ES cell colonies consisted of a mixture of cells that have either retained or lost the conditional allele. We demonstrate that these mixed ES cell clones can be directly used for generation of chimeras that give rise to offspring with conditional or knock-out alleles simultaneously. This strategy shortens the time and reduces the number of germline transmission events to generate genetically modified mice.

译文

功能丧失的小鼠模型包括敲除小鼠(其中种系中的基因缺失)和条件性敲除小鼠,其在定义的组织中体细胞性等位基因缺失。两种类型的小鼠都用于体内基因功能的综合研究。在这里,我们描述了使用单个胚胎干(ES)细胞克隆同时生成具有转录因子fra-2(Fos相关抗原2)的条件或敲除等位基因的小鼠的简单方法。用Cre-重组酶表达质粒瞬时转染带有fra-2等位基因的ES细胞,并以低密度铺板。产生的大多数ES细胞集落由保留或丢失条件等位基因的细胞混合物组成。我们证明,这些混合的ES细胞克隆可直接用于产生嵌合体,这些嵌合体同时产生条件等位基因或敲除等位基因。该策略缩短了时间并减少了产生遗传修饰小鼠的种系传播事件的数量。

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