A Novel Co-immunoprecipitation Protocol Based on Protoplast Transient Gene Expression for Studying Protein-protein Interactions in Rice
A Novel Co-immunoprecipitation Protocol Based on Protoplast Transient Gene Expression for Studying Protein-protein Interactions in Rice
复制标题
一种基于原生质体瞬时基因表达的新型免疫共沉淀方案，用于研究水稻中蛋白质-蛋白质相互作用
DOI：
10.1007/s11105-013-0633-9
复制
发表时间：
2014-02-01
影响因子：
2.1
通讯作者：
Zhuang, Chu Xiong
中科院分区：
文献类型：
--
作者：
Yang, Jia Wei;Fu, Jia Xin;Zhuang, Chu Xiong
蛋白质-蛋白质相互作用构成了协调不同细胞功能的调节网络。免疫共沉淀（co-IP）是一种广泛使用的和有效的技术，研究蛋白质-蛋白质相互作用的活细胞。然而，制备高度特异性抗体的时间和成本是与该技术相关的主要缺点。在本研究中，一个co-IP系统开发的基础上改进的原生质体瞬时表达系统，通过使用商业上可获得的抗体检测蛋白质-蛋白质相互作用。这种co-IP系统消除了对特异性抗体制备和转基因植物生产的需要。以水稻绿色幼苗叶鞘为材料，建立了原生质体瞬时表达系统，并进行了质粒共转化。本研究建立的瞬时表达系统适用于亚细胞定位和蛋白质检测。这项工作提供了一个快速，可靠，具有成本效益的系统，研究瞬时基因表达，蛋白质亚细胞定位，并在体内蛋白质-蛋白质相互作用的表征。
Protein-protein interactions constitute the regulatory network that coordinates diverse cellular functions. Co-immunoprecipitation (co-IP) is a widely used and effective technique to study protein-protein interactions in living cells. However, the time and cost for the preparation of a highly specific antibody is the major disadvantage associated with this technique. In the present study, a co-IP system was developed to detect protein-protein interactions based on an improved protoplast transient expression system by using commercially available antibodies. This co-IP system eliminates the need for specific antibody preparation and transgenic plant production. Leaf sheaths of rice green seedlings were used for the protoplast transient expression system which demonstrated high transformation and co-transformation efficiencies of plasmids. The transient expression system developed by this study is suitable for subcellular localization and protein detection. This work provides a rapid, reliable, and cost-effective system to study transient gene expression, protein subcellular localization, and characterization of protein-protein interactions in vivo.