Fig. 2. Timing of TagA expression. (A) RNase protection assays were performed with total RNA collected from wild-type (upper panel) or tagA mutant (lower panel) cells. Lanes are: P, riboprobe without RNase treatment (1/10 the input for other lanes), Y, RNase digestion of probe incubated with yeast RNA, and (R) RNase digestion of the probe without RNA added, or (0-24 h) after hybridization to RNA samples collected across the 24 hours of development. (B) Western blot stained with a TagA antibody detects a protein of an apparent molecular mass of 190 kDa (arrow). Equal amounts of protein (10 µg) from vegetative (0 hours) or developing (2-24 hours) wild-type cells (Ax4) were loaded in each lane, along with molecular mass standards (MM). The tagA mutant and rescued mutant (tagA^-[tagA/tagA]) samples were mixtures of all vegetative and developmental time points. The amount of protein loaded in these lanes were equal (1x) or twofold (2x) the amounts of the developmental samples.