Abstract
Immunoprecipitation of mRNA-protein complexes is a method that can be used to study RNA binding protein (RBP)–RNA interactions. In this protocol, an antibody targeting an RBP of interest is used to immunoprecipitate the RBP and any interacting molecules from a cell lysate. Reverse transcription followed by PCR is then used to identify individual mRNAs isolated with the RBP. This method focuses on examining an association between a specific RBP-mRNA complex, and it is best suited for a small scale screening of known or putative binding partners. It can also be used as a second, independent method to verify RBP-mRNA interactions discovered through more universal screening techniques. We describe the immunoprecipitation protocol in practical detail and discuss variations of the method as well as issues associated with it. The procedure takes three days to complete.
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References
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Acknowledgements
We thank M. Maronski for help with culturing the neurons. This work was funded by grants AG9900 and MH58561 from the NIH to JE and the Swedish Science Foundation Grants Med and NT, and European Community Grant QLRT-2001-01989 to U.L.
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Peritz, T., Zeng, F., Kannanayakal, T. et al. Immunoprecipitation of mRNA-protein complexes. Nat Protoc 1, 577–580 (2006). https://doi.org/10.1038/nprot.2006.82
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DOI: https://doi.org/10.1038/nprot.2006.82
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