UW-Stout/Ethanol FA22

Yeast

Yeast Ethanol Stress Experiment

 Introduction
   We will add a sample of ethanol solution to a yeast culture and measure the growth afterward. This experiment is to determine what amounts of ethanol solution are best for the tress experiment on our class knockout genes.

Materials 

• Corning COSTAR 96-well clear flat-bottom assay plate • Wild-type yeast in 2x synthetic complete media at an OD600 of 0.1-0.2 • Ethanol concertation of 1M (labeled ETH01M)

Procedure 

1. Make the ethanol solution by add 942µl of water and 58µl of 100% ethanol 2. Collect well a clear flat-bottom assay plate 3. Vortex yeast culture for about 5 seconds 4. Put 50µl of the yeast culture into all 12 wells. 5. In the 1st well add 1µl of your ethanol solution and 49µl of H2O 6. In the 2nd well add 2µl of your ethanol solution and 48µl of H2O 7. In the 3rd well add 3µl of your ethanol solution and 47µl of H2O 8. In the 4th well add 4µl of your ethanol solution and 46µl of H2O 9. In the 5th well add 5µl of your ethanol solution and 45µl of H2O 10. In the 6th well add 8µl of your ethanol solution and 42µl of H2O 11. In the 7th well add 10µl of your ethanol solution and 40µl of H2O 12. In the 8th well add 20µl of your ethanol solution and 30µl of H2O 13. In the 9th well add 27µl of your ethanol solution and 23µl of H2O 14. In the 10th well add 35µl of your ethanol solution and 15µl of H2O 15. In the 11th well add 41µl of your ethanol solution and 9µl of H2O 16. In the 12th well add 50µl of your ethanol solution and 0µl of H2O 17. In E column 12th well add 0µl of your ethanol solution and 50µl of H2O 18. Put your well clear flat-bottom assay plate into the incubator and let sit. 19. Wait and observe the growth or death