Abstract
Protein–protein interactions constitute the regulatory network that coordinates diverse cellular functions. Co-immunoprecipitation (co-IP) is a widely used and effective technique to study protein–protein interactions in living cells. However, the time and cost for the preparation of a highly specific antibody is the major disadvantage associated with this technique. In the present study, a co-IP system was developed to detect protein–protein interactions based on an improved protoplast transient expression system by using commercially available antibodies. This co-IP system eliminates the need for specific antibody preparation and transgenic plant production. Leaf sheaths of rice green seedlings were used for the protoplast transient expression system which demonstrated high transformation and co-transformation efficiencies of plasmids. The transient expression system developed by this study is suitable for subcellular localization and protein detection. This work provides a rapid, reliable, and cost-effective system to study transient gene expression, protein subcellular localization, and characterization of protein–protein interactions in vivo.
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Acknowledgments
This work was supported by the National Basic Research Program of China (2013CBA01401), Genetically Modified Organisms Breeding Major Projects (2013ZX08001-004), the National Natural Science Foundation of China (No. 30800596 and No. 31200191) and China Postdoctoral Science Foundation (No. 20110490897). We thank Professor Li-Zhen Tao in South China Agricultural University for help with fluorescent microscope analysis; Professor Zheng-Hui He in San Francisco State University and Shi-Jiang Zhu in South China Agricultural University for the comments on the manuscript and Cole Johnson in University of Michigan for writing assistance.
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J.W. Yang and J.X. Fu both authors contributed equally to this work.
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Yang, J.W., Fu, J.X., Li, J. et al. A Novel Co-immunoprecipitation Protocol Based on Protoplast Transient Gene Expression for Studying Protein–protein Interactions in Rice. Plant Mol Biol Rep 32, 153–161 (2014). https://doi.org/10.1007/s11105-013-0633-9
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DOI: https://doi.org/10.1007/s11105-013-0633-9