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Simple Nuclei Preparation and Co-immunoprecipitation Procedures for Studying Protein Abundance and Interactions in Plant Circadian Time Courses

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Plant Circadian Networks

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2398))

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Abstract

The plant circadian clock regulates multiple developmental and physiological events that occur at specific times and seasons. As many of the currently known clock proteins and clock-associated regulators are transcription factors, analyzing molecular events in the nuclei is crucial. In addition, long-time course analyses of protein abundance and interactions are often required to assess the role of the circadian clock on clock-regulated phenomena. Here we introduce a simple procedure to prepare nuclear-enriched tissues, which we routinely use to study time-resolved accumulation changes in low-abundance nuclear proteins (i.e., transcription factors). In addition to measuring changes in abundance, investigating the protein-protein interaction dynamics at specific times of day or under certain environmental conditions is needed for plant chronobiology studies. Therefore, we also present our co-immunoprecipitation method for studying diurnal/circadian protein-protein interactions, tailored to nuclear-localized proteins in Arabidopsis and tobacco.

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Acknowledgments

This work was supported by NIH grant (GM079712) to TI, the National Research Foundation of Korea (NRF) grant (NRF-2020R1A2C1014655) to Y.H.S., JSPS KAKENHI (19K16170), Grant-in-Aid for Scientific Research on Innovative Areas (JP19H04866), Start-up Grant for Women Scientists from NAIST, and Lifescience Grant from Takeda Science Foundation to A.K.

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Correspondence to Takato Imaizumi .

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Kubota, A., Song, Y.H., Imaizumi, T. (2022). Simple Nuclei Preparation and Co-immunoprecipitation Procedures for Studying Protein Abundance and Interactions in Plant Circadian Time Courses. In: Staiger, D., Davis, S., Davis, A.M. (eds) Plant Circadian Networks. Methods in Molecular Biology, vol 2398. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1912-4_11

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  • DOI: https://doi.org/10.1007/978-1-0716-1912-4_11

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1911-7

  • Online ISBN: 978-1-0716-1912-4

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