Abstract
Stable isotope labeling by amino acids combined with mass spectrometry is a widely used methodology for measuring relative changes in protein and phosphorylation levels at a global level. We have applied this method to the model organism Caenorhabditis elegans in combination with RNAi-mediated gene knockdown by feeding the nematode on pre-labeled lysine auxotroph Escherichia coli. In this chapter, we describe in details the generation of the E. coli strain, incorporation of heavy isotope-labeled lysine in C. elegans, and the procedure for a comprehensive global phosphoproteomic experiment.
Julius Fredens and Kasper Engholm-Kelle have equally contributed to this Chapter.
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Acknowledgements
This work was supported by The Lundbeck Foundation and The Danish Council for Independent Research, Natural Sciences.
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Fredens, J., Engholm-Keller, K., Møller-Jensen, J., Larsen, M.R., Færgeman, N.J. (2014). Identification of Novel Protein Functions and Signaling Mechanisms by Genetics and Quantitative Phosphoproteomics in Caenorhabditis elegans . In: Warscheid, B. (eds) Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC). Methods in Molecular Biology, vol 1188. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1142-4_9
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DOI: https://doi.org/10.1007/978-1-4939-1142-4_9
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