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Monitoring of Ubiquitin-proteasome Activity in Living Cells Using a Degron Dgn-destabilized Green Fluorescent Protein GFP-based Reporter Protein Video (Video) | JoVE

DOI :

10.3791/3327-v

November 10th, 2012

November 10th, 2012

16,484 Views

1Molecular and Cell Biology, Institute for Biomedical Aging Research, 2Department of Gerontology and Geriatrics, Netherlands Consortium for Healthy Aging, Leiden University Medical Center

A method to monitor ubiquitin-proteasome activity in living cells is described. A degron-destabilized GFP- (GFP-dgn) and a stable GFP-dgnFS fusion protein are generated and transduced into the cell using a lentiviral expression vector. This technique allows to generate a stable GFP-dgn/GFP-dgnFS expressing cell line in which ubiquitin-proteasome activity can be easily assessed using epifluorescence or flow cytometry.

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