Graduate School of Pharmaceutical Sciences, Nagoya University, Nagoya, Japan.
Graduate School of Agriculture, Kyoto University, Kyoto, Japan.
Biosci Biotechnol Biochem. 2023 Oct 25;87(11):1285-1294. doi: 10.1093/bbb/zbad116.
Deletion of gene expression in the target tissues and cells is an effective strategy for elucidating the physiological functions of the protein of interest. For tissue-specific and/or inducible gene deletion, the Cre-loxP system has been widely used in various model organisms including medaka (Oryzias latipes). The epithelium is the key tissue, locating at the outermost area and playing a role in barrier to external stimuli. Despite a large genetic toolbox developed in medaka, there is no available Cre-driver line that works in an epithelium-specific manner. Here, we established epithelium-specific Cre-driver lines in medaka using a homology-directed repair mediated knock-in approach with CRISPR/Cas9, targeting each of periplakin and keratin genes. We show that Cre-recombinase is expressed exclusively in the epithelium in the knock-in lines and that it efficiently and specifically induces recombination in the tissues. These Cre-driver lines are useful for studying the functions of proteins expressed in the epithelium.
在靶组织和细胞中删除基因表达是阐明目的蛋白生理功能的有效策略。对于组织特异性和/或诱导性基因缺失，Cre-loxP 系统已广泛应用于包括斑马鱼（Oryzias latipes）在内的多种模式生物中。上皮组织是关键组织，位于最外层区域，在抵御外部刺激方面发挥作用。尽管在斑马鱼中开发了大量的遗传工具，但没有可用的以上皮组织特异性方式工作的 Cre 驱动线。在这里，我们使用 CRISPR/Cas9 介导的同源定向修复基因敲入方法，针对整联蛋白和角蛋白基因，在斑马鱼中建立了上皮组织特异性 Cre 驱动线。我们表明，在敲入系中，Cre 重组酶仅在上皮组织中表达，并且它能有效地、特异性地诱导组织中的重组。这些 Cre 驱动线可用于研究在上皮组织中表达的蛋白的功能。
