FIGURE 2.

Expression of the ADH6 and ADH7 genes under vanillin stress. (A) The mRNA levels of ADH6 and ADH7 were analyzed by qRT-PCR. Cells in an exponential phase of growth were treated with vanillin (6–15 mM) for 60 min. To compare mRNA expression levels, the mRNA level of each gene was normalized to that of ACT1. The mRNA level in cells without vanillin treatment was considered 100%. Data are shown as mean ± SD of three independent experiments. (B,C) Levels of Adh6 and Adh7 protein expression were determined by western blot analysis using an anti-FLAG antibody. Cells carrying a FLAG-tagged chromosomal copy of ADH6 or ADH7 gene were treated with the indicated concentrations of vanillin for 60 min (B), or treated with 8 mM vanillin for the indicated periods of time (C). Pgk1 was used as a loading control. Protein levels of Adh6 and Adh7 were normalized to that of Pgk1 using ImageJ, and the intensity of Pgk1 band of each lane was considered 100%. Data are shown as the mean ± SD of three independent experiments.