All strains were maintained in cryo-culture at –80°C in 25% glycerol and passaged in YPD. For solid medium, 2% agar was used. Strains that were auxotrophic for uridine were grown with 80 mg/L uridine added to the growth medium. When indicated, strains were grown in the presence of DOX (Doxycycline Hydrochloride, DB0889, BioBasic, Markham, ON, Canada) dissolved in water. For experiments to repress gene expression, unless indicated otherwise, overnights of relevant strains and controls were subcultured in the absence or presence of 0.5 μg/mL DOX and grown overnight. The cells were subcultured into 5 μg/mL DOX and grown to mid-log phase for qRT-PCR experiments and western blot analysis.
For heat shock experiments, overnights of relevant strains were grown in YPD at 30°C and then diluted to an OD600 of 0.1 and subcultured for 4 hours to mid-log phase before 10 minutes of heat shock at the indicated temperatures.
Individual strains are listed inS5 Table . Plasmids used for strain construction are included in S6 Table . All primer sequences are included in S7 Table .
For heat shock experiments, overnights of relevant strains were grown in YPD at 30°C and then diluted to an OD600 of 0.1 and subcultured for 4 hours to mid-log phase before 10 minutes of heat shock at the indicated temperatures.
Individual strains are listed in