一种蛋白质组学方案，用于鉴定哺乳动物细胞中依赖于特定基因的刺激诱导结合伙伴。
A proteomics protocol to identify stimulation-induced binding partners dependent on a specific gene in mammalian cells.
机构信息
State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, China.
The Innovation of Seed Design, Chinese Academy of Sciences, Wuhan 430072, China.
出版信息
STAR Protoc. 2021 Nov 13;2(4):100962. doi: 10.1016/j.xpro.2021.100962. eCollection 2021 Dec 17.
Some protein-protein interactions are induced by different kinds of stimulation and are dependent on specific genes. To identify these interaction partners, we present a protocol which utilizes affinity purification of Flag-tagged protein complexes followed by mass-spectrometry-based proteomics to compare stimulation-induced interactomes between wild-type and CRISPR-Cas9-mediated knockout cells. The candidates of interest are identified using bioinformatic analyses and verified by biochemical approaches. This protocol is highly versatile and applies to a variety of cells and different types of stimulation. For complete details on the use and execution of this protocol, please refer to (Zhu et al., 2021).
有些蛋白质-蛋白质相互作用是由不同的刺激诱导的，并依赖于特定的基因。为了鉴定这些相互作用伙伴，我们提出了一种方案，该方案利用 Flag 标记的蛋白复合物的亲和纯化，然后进行基于质谱的蛋白质组学分析，以比较野生型和 CRISPR-Cas9 介导的基因敲除细胞之间的刺激诱导的相互作用组。使用生物信息学分析来鉴定感兴趣的候选物，并通过生化方法进行验证。该方案具有高度的通用性，适用于各种细胞和不同类型的刺激。有关该方案的使用和执行的完整详细信息，请参考（Zhu 等人，2021 年）。
Zotero 插件