- Corning COSTAR 96-well clear flat-bottom assay plate
- Falcon round-bottom polypropylene tubes
- YPD media, both broth and 2% agar plates
- Glycerol stocks of knockout strains
- Incubator set to 30°C.
- New Brunswick TC-7 Tissue Culture Roller Drum
- Molecular Devices SpectraMax Plus 384 Microplate Reader
- Three days before the experiment, streak knockout strains onto YPD plates.
- The morning before the experiment, pick a colony from each strain into 5 ml YPD broth in a disposable test-tube. Incubate on the roller drum at 30°C for 8-10 hours.
- The evening before the experiment, transfer 20 ul of each strain into 5 ml of fresh YPD broth. Incubate on a roller drum at 30°C overnight.
- The morning of the experiment, dilute each culture to an OD600 of 0.1-0.2 in YPD broth. Return to the drum roller until ready to test.
- Transfer 50 µl of yeast culture and 50 µl of sterile water to a well in the assay plate.
- Set up the plate reader as follows:
- Temperature: 30°C
- Mode: Kinetic
- Read: 600 nm
- Interval: 5 minutes
- Total run time: 24 hours
- Shake before read: 30 seconds
- Transfer the assay plate to the reader and run for 24 hours.