Profiling post-transcriptionally networked mRNA subsets using RIP-Chip and RIP-Seq
Profiling post-transcriptionally networked mRNA subsets using RIP-Chip and RIP-Seq
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DOI：
10.1016/j.ymeth.2013.11.001
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发表时间：
2014-05-01
期刊：
影响因子：
4.8
通讯作者：
Tenenbaum, Scott A.
中科院分区：
文献类型：
--
作者：
Jayaseelan, Sabarinath;Doyle, Francis;Tenenbaum, Scott A.
信使RNA的转录后调控有助于基因表达的许多方面。这一调控水平的关键组成部分是rna结合蛋白（rbp）与其相关靶mRNA的相互作用。剪接、稳定性、定位、翻译效率和密码子的交替使用只是rbp调控的一些转录后过程。我们对这些过程的理解的核心是需要表征RBP-mRNA关联网络，并创建这种功能性转录后调控系统的图谱。在这里，我们提供了一种详细的方法来分离mRNA，使用RBP免疫沉淀（RIP）作为主要的划分方法，然后是微阵列（Chip）或下一代测序（NGS）分析。我们通过使用特异性抗体来靶向rbp以捕获相关的RNA货物来做到这一点。RIP-Chip/Seq已被证明是一种通用的基因组技术，已被广泛用于研究内源性RBP-RNA关联。Elsevier Inc.出版。
Post-transcriptional regulation of messenger RNA contributes to numerous aspects of gene expression. The key component to this level of regulation is the interaction of RNA-binding proteins (RBPs) and their associated target mRNA. Splicing, stability, localization, translational efficiency, and alternate codon use are just some of the post-transcriptional processes regulated by RBPs. Central to our understanding of these processes is the need to characterize the network of RBP-mRNA associations and create a map of this functional post-transcriptional regulatory system. Here we provide a detailed methodology for mRNA isolation using RBP immunoprecipitation (RIP) as a primary partitioning approach followed by microarray (Chip) or next generation sequencing (NGS) analysis. We do this by using specific antibodies to target RBPs for the capture of associated RNA cargo. RIP-Chip/Seq has proven to be is a versatile, genomic technique that has been widely used to study endogenous RBP-RNA associations. Published by Elsevier Inc.