Antisense-mediated affinity purification of dengue virus ribonucleoprotein complexes from infected cells.
Antisense-mediated affinity purification of dengue virus ribonucleoprotein complexes from infected cells.
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从受感染细胞中反义介导的登革热病毒核糖核蛋白复合物亲和纯化。
DOI：
10.1016/j.ymeth.2015.08.008
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发表时间：
2015
期刊：
影响因子：
--
通讯作者：
Bradrick,SheltonS
中科院分区：
文献类型：
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作者：
Phillips,StaciaL;Garcia-Blanco,MarianoA;Bradrick,SheltonS
鉴定在感染过程中与病毒 RNA 分子物理结合的 RNA 结合蛋白可以深入了解 RNA 病毒复制的分子机制。直到最近，这种 RNA-蛋白质相互作用主要通过使用体外测定来识别，但可能无法准确反映活细胞环境中发生的关联。在这里，我们描述了一种使用体内交联然后进行反义介导的亲和纯化来特异性亲和纯化登革热病毒RNA和相关蛋白的方法。使用这种方法与病毒 RNA 特异性共纯化的 RNA 结合蛋白可以通过质谱法进行鉴定。该策略有可能适用于任何感兴趣的病毒 RNA 种类的纯化。
The identification of RNA-binding proteins that physically associate with viral RNA molecules during infection can provide insight into the molecular mechanisms of RNA virus replication. Until recently, such RNA–protein interactions have been identified predominantly with the use ofin vitroassays that may not accurately reflect associations that occur in the context of a living cell. Here we describe a method for the specific affinity purification of dengue virus RNA and associated proteins usingin vivocross-linking followed by antisense-mediated affinity purification. RNA-binding proteins that specifically co-purify with viral RNA using this method can be identifieden masseby mass spectrometry. This strategy can potentially be adapted to the purification of any viral RNA species of interest.