Label-free bottom-up shotgun MS-based proteomics is an extremely powerful and simple tool to provide high quality quantitative analyses of the yeast proteome with only microgram amounts of total protein. Although the experimental design of this approach is rather straightforward and does not require the modification of growth conditions, proteins or peptides, several factors must be taken into account to benefit fully from the power of this method. Key factors include the choice of an appropriate method for the preparation of protein extracts, careful evaluation of the instrument design and available analytical capabilities, the choice of the quantification method (intensity-based vs. spectral count), and the proper manipulation of the selected quantification algorithm. The elaboration of this robust workflow for data acquisition, processing, and analysis provides unprecedented insight into the dynamics of the yeast proteome.
Keywords: Label-free quantification; Mass spectrometry; Yeast proteomics.