Fan Baochang, Ni Peng, Kao C Cheng
Department of Molecular and Cellular Biochemistry, Indiana University, 212 S. Hawthorne St., 201A Simon Hall, Bloomington, IN, 47405, USA.
Methods Mol Biol. 2015;1297:213-24. doi: 10.1007/978-1-4939-2562-9_15.
RNA nanotechnology often involves protein-RNA complexes that require significant understanding of how the proteins and RNAs contact each other. The CLIP-Seq (cross-linking immunoprecipitation, and DNA sequencing) protocol can be used to probe the RNA molecules that interact with proteins. We have optimized the procedures for RNA fragmentation, immunoprecipitation, and library construction in CLIP-Seq to map the interactions between the RNA and the capsid of a simple positive-strand RNA virus. The results show that distinct portions of the viral RNA contact the capsid. The protocol should be applicable to other RNA virions and also RNA-protein nanoparticles.
RNA纳米技术通常涉及蛋白质-RNA复合物，这需要深入了解蛋白质和RNA如何相互接触。CLIP-Seq（交联免疫沉淀和DNA测序）方案可用于探测与蛋白质相互作用的RNA分子。我们优化了CLIP-Seq中RNA片段化、免疫沉淀和文库构建的程序，以绘制一种简单正链RNA病毒的RNA与衣壳之间的相互作用。结果表明，病毒RNA的不同部分与衣壳接触。该方案应适用于其他RNA病毒粒子以及RNA-蛋白质纳米颗粒。
