Abstract
Ubiquitin is a small protein commonly used as a signal molecule which upon attachment to the proteins affects their function and their fate in the cells. For example, it can be used as a degradation marker by the cell. Ubiquitin plays a significant role in regulation of numerous cellular processes. Therefore, monitoring of ubiquitin-dependent proteolysis can provide important information. Here, we describe construction of YFP-based proteasome substrates containing modified ubiquitin and the protocol for their transient expression in plant cells for functional analysis of the ubiquitin/proteasome system. To facilitate further subcloning all plasmids generated by us are based on the Gateway® Cloning Technology and are compatible with the Gateway® destination vectors.
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Acknowledgments
This work was supported by the Ministry of Science and Higher Education in frame of the project No W16/7.PR/2011 realized in the years 2011–2015 and the National Science Centre (grant No 2012/05/N/NZ1/00699 realized in the years 2013–2014). KZ-R was also supported by the European Union Mazovia Fellowship “Development of science—development of the region—scholarship support for Mazovias’ Ph.D.’s”.
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Zientara-Rytter, K., Sirko, A. (2016). Fluorescent Reporters for Ubiquitin-Dependent Proteolysis in Plants. In: Lois, L., Matthiesen, R. (eds) Plant Proteostasis. Methods in Molecular Biology, vol 1450. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3759-2_5
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DOI: https://doi.org/10.1007/978-1-4939-3759-2_5
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Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3757-8
Online ISBN: 978-1-4939-3759-2
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