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May 24, 2014
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Here, we describe a methodology to perform a targeted siRNA “ubiquitome” screen to identify novel ubiquitin and ubiquitin-like regulators of the HIF1A-mediated cellular response to hypoxia. This can be adapted to any biological pathway where a robust read out of reporter activity is available.
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SiRNA ScreeningUbiquitinUbiquitin-like SystemBiological PathwaysMammalian CellsPost-translational ModificationUbiquitomeHIF1AHypoxia ResponseTranscription-based Luciferase ReporterAssay DevelopmentPrimary ScreeningSecondary ScreeningTertiary/deconvolution ScreeningTargeted SiRNA LibraryOff-target Effects
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Bett, J. S., Ibrahim, A. F. M., Garg, A. K., Rocha, S., Hay, R. T. siRNA Screening to Identify Ubiquitin and Ubiquitin-like System Regulators of Biological Pathways in Cultured Mammalian Cells. J. Vis. Exp. (87), e51572, doi:10.3791/51572 (2014).
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