Immunopurification and Analysis of Protein and RNA Components of mRNP in Mammalian Cells
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Last Update: 2020-12-20
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Source: Internet
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Author: User
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We describe a basic, fast, and reliable technique to isolate and characterize ribonucleoprotein (RNP) using antibody to aconstituent protein. The antibody serves to immunopurify RNP from total cells or nuclear and cytoplasmic cell fractions underconditions that promote RNP integrity. The presence of other RNP proteins as well as transcripts can then be analyzed by Westernblotting and reverse transcription polymerase chain reaction, respectively. RNase treatment before immunopurification canbe used to assess the dependence of protein-protein interactions on RNA. We also describe a modification using β-mercaptoethanolthat facilitates analyzing proteins that comigrate with antibody light or heavy chains.
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