Figure 4
From: C-terminal processing of yeast Spt7 occurs in the absence of functional SAGA complex
Time course of the induction and decay of Spt7 forms. A. CY1811 containing YIplac211-GAL-Flag3-SPT7 was grown in minimal media containing 2% raffinose then diluted 1:5 into YP media containing 2% glucose (lane 2) or 2%galactose (lane 3) and grown for 30 minutes. Extracts were prepared by glass bead disruption and 100 μg separated by SDS-PAGE (5%). Flag3-Spt7 was detected by Western blotting with anti-Flag antibody. Lane one contains 100 μg of protein extract prepared after growth of BY4741 in galactose-containing media. B. CY1811 containing YIplac211-GAL – Flag3-SPT7 was grown in raffinose. Expression of Spt7 was induced by the addition of galactose then inhibited by the addition of YP media containing 2% glucose. Cells were grown for 90 minutes which was empirically determined as time 0. Additional equal volume samples were taken at time 0, 30, 60, 120 and 240 minutes (lanes 2–7). Protein extracts were prepared and equal volumes separated by SDS-Page (5%). Flag3-Spt7 was detected by Western blotting (top panel) or stained with Coomassie Brilliant Blue (bottom panel).