Mabuchi Naoto, Masuyama Kaoru, Ohno Mutsuhito
Institute for Virus Research, Kyoto University, Japan.
Methods Mol Biol. 2008;488:257-65. doi: 10.1007/978-1-60327-475-3_18.
Results obtained from in vitro experiments often need to be confirmed by in vivo experiments. The study of RNA-protein interactions is no exception. Information on RNA-protein complex formation in the cell is important for understanding the mechanisms of cellular RNA metabolism such as RNA processing and transport. For such purposes, Xenopus oocytes are extremely useful cells thanks to their large size. Interactions of microinjected proteins and RNAs with their binding partners can be examined easily by immunoprecipitation experiments with nuclear or cytoplasmic fractions from microinjected Xenopus oocytes. We describe a method to study how RNAs that have been microinjected into the nucleus of Xenopus oocytes are assembled into complexes with specific endogenous proteins.
体外实验获得的结果通常需要通过体内实验来证实。RNA-蛋白质相互作用的研究也不例外。细胞中RNA-蛋白质复合物形成的信息对于理解细胞RNA代谢机制(如RNA加工和转运)很重要。出于这些目的,非洲爪蟾卵母细胞因其体积大而成为极其有用的细胞。通过对显微注射的非洲爪蟾卵母细胞的细胞核或细胞质组分进行免疫沉淀实验,可以轻松检测显微注射的蛋白质和RNA与其结合伴侣的相互作用。我们描述了一种研究显微注射到非洲爪蟾卵母细胞核中的RNA如何与特定内源性蛋白质组装成复合物的方法。