UW-Stout/Heat Shock SP21

Preliminaries: None

Calibration Experiment Materials: • Distilled H2O • Wild-type yeast cells • P20 and P200 micro pipettors • PCR test tubes (mini) • Glass beads • 15 agar filled petri dishes

Calibration Experiment Procedure:

1. Vortex the yeast culture briefly to resuspend the yeast pellet. 2. Set up 5 different PCR test tubes with 50 ul of wild-strain yeast cells and 50 ul of distilled water in each. 3. Label each tube “A, B, C, D, E”. 4. Insert test tube A in the thermocycler for exactly 30 minutes at 48°C. 5. After test tube A is done cycling, place test tube B in the thermal cycler for exactly 15 minutes at 48° C. 6. After test tube B is done cycling, place test tube C in the thermal cycler for exactly 30 minutes at 48° C. 7. After test tube C is done cycling, place test tube D in the thermal cycler for exactly 45 minutes at 48° C. 8. After test tube D is done cycling, place test tube E in the thermal cycler for exactly 60 minutes at 48° C. 9. Carry on and analyze/count the DNA colonies in the wells.

1 2 3 4 5 PCR Tubes with Volume 50 ul yeast / 50 ul H2O 50 ul yeast 50 ul water 50 ul yeast 50 ul water 50 ul yeast 50 ul water 50 ul yeast 50 ul water 50 ul yeast 50 ul water Time (Minutes) 5 minutes 15 minutes 30 minutes 45 minutes 60 minutes

Knockout Experiment Materials: