Comparison of 2-Dimensioal protein gels from wild type and mutant yeast strains as analyzed by 2 different instrument systems

Proteomic analysis of biological samples is increasing in its importance for various research applications. Part and parcel of this increase in interest is an influx of commercial formats available for utilizing this technology. Because cost and application are generally limiting factors when choosing among the various instrumentation available, one challenge for a core facility is then to choose among these available formats one which can fulfill the needs of both client and facility. To this end a robust comparative analysis of competing platforms would be beneficial. Our experimental models were a tRNA methyltransferase (Trm10) deletion strain of Saccharomyces cerevisiae and its otherwise isogenic wildtype parent strain. The trm10Δ strain is hypersensitive to the presence of the anti-tumor agent 5fluorouracil (5-FU), but the biological basis for this hypersensitivity is not known, and identification of proteomic changes that result from growth in the presence of 5-FU could provide important clues to the mechanism of 5-FU action in cells. Following overnight treatment with a sub-lethal dose of 5-FU, total soluble protein from both strains was isolated and a portion was labeled using the GE Healthcare DIGE labeling system and subjected to separation by 2D PAGE. The resulting gels were (1) subsequently visualized on the Bio Rad Versa Doc 4000MP Imaging platform and analyzed with PD Quest and (2) imaged with the GE Healthcare Typhoon system and analyzed with DeCyder for comparison of the platforms. The same protein isolates were also separated by 2D PAGE and stained with SYPRO Ruby protein stain and subsequently visualized on both imaging platforms. Images were analyzed using the GE Healthcare DeCyder and Bio-Rad PD Quest analysis software packages as a further comparison group.