Jayaseelan Sabarinath, Doyle Francis, Currenti Salvatore, Tenenbaum Scott A
Department of Nanobioscience, University at Albany, Albany, NY, USA.
Methods Mol Biol. 2011;714:407-22. doi: 10.1007/978-1-61779-005-8_25.
A detailed understanding of post-transcriptional gene expression is necessary to correlate the different elements involved in the many levels of RNA-protein interactions that are needed to coordinate the cellular biomolecular machinery. The profile of mRNA, a major component of this machinery, can be examined after isolation from specific RNA-binding proteins (RBPs). RIP-Chip or ribonomic profiling is a versatilein vivo technique that has been widely used to study post-transcriptional gene regulation and the localization of mRNA. Here we elaborately detail the methodology for mRNA isolation using RBP immunoprecipitation (RIP) as a primary approach. Specific antibodies are used to target RBPs, which are then used to capture the associated mRNA.
要将协调细胞生物分子机制所需的RNA-蛋白质相互作用多个层面中涉及的不同元件关联起来,就必须深入了解转录后基因表达。该机制的一个主要组成部分——信使核糖核酸(mRNA)的概况,可在从特定RNA结合蛋白(RBP)中分离出来后进行检测。RIP-Chip或核糖核酸分析是一种通用的体内技术,已被广泛用于研究转录后基因调控和mRNA的定位。在此,我们详细阐述了以RBP免疫沉淀(RIP)作为主要方法分离mRNA的方法。使用特异性抗体靶向RBP,然后用其捕获相关的mRNA。