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Cell Line Nucleofector Kit V

Company: Amaxa
Catalog#: VCA-1003
Other protocol()

Ciliary Assembly/Disassembly Assay in Non-transformed Cell Lines
[Abstract]  The primary cilium is a non-motile sensory organelle whose assembly and disassembly are closely associated with cell cycle progression. The primary cilium is elongated from the basal body in quiescent cells and is resorbed as the cells re-enter the cell cycle. Dysregulation of ciliary dynamics has been linked with ciliopathies and other human diseases. The in vitro serum-stimulated ciliary assembly/disassembly assay has gained popularity in addressing the functions of the protein-of-interest in ciliary dynamics. Here, we describe a well-tested protocol for transfecting human retinal pigment epithelial cells (RPE-1) and performing ciliary assembly/disassembly assays on the transfected cells. [摘要]  主要纤毛是一种非运动感觉细胞器,其装配和拆卸与细胞周期进程密切相关。 初级纤毛在静止细胞中从基体拉长并随着细胞重新进入细胞周期而被吸收。 睫状动力失调与纤毛病和其他人类疾病有关。 体外血清刺激的睫状体装配/分解测定已经在解决睫状动力学中感兴趣的蛋白质的功能方面受到欢迎。 在这里,我们描述了转染人视网膜色素上皮细胞(RPE-1)和对转染细胞进行睫状体装配/分解测定的充分测试的方案。

【背景】初级纤毛是毛发样感觉细胞器,其在G 0 / G 1期出现,并且在细胞周期的S期之前分解(Tucker等, et al。,1979)。先前的研究已经证实,某些未转化的细胞类型(即,甚至是RPE-1细胞,3T3成纤维细胞和小鼠胚胎成纤维细胞[MEFs])可以被饿死以诱导静止和睫状体形成。随后的血清再次添加触发双相睫状体吸收,其在刺激后2小时和24小时达到峰值(Tucker等人,1979; Li等人,2011) 。该现象为文献中常用的血清刺激的睫状体组装/分解测定奠定了基础,以鉴定参与睫状体组装和拆卸的蛋白质(Pugacheva等人,2007; ...