UW Stout/Sucrose Fermentation SP22

Introduction

The purpose of this protocol is to test if knockout yeast strains undergo fermentation, and if they do how much ethanol is produced. This will tell us if the gene that was knocked out plays a role in fermentation.

Materials

• 1 ml Yeast Culture (Approximately 1 million cells)
  • You may need to dilute with sterile H2O to approximate the 1 million cells. Keep total volume at 1 ml.
• PBS Buffer Solution
• Yeast Growth Solution
  • 0.85g YNB, 2.5 Ammonium Sulfate, 0.32g CSM, 5ml 100x Leucine, 100x Histine, 100x Uracil, 435 ml DI H2O
• 40% Sucrose Solution

Procedure

Set up yeast

1. Collect 1 ml of yeast solution
2. Centrifuge for 3 minutes at 1000x gravity
3. Remove supernatant
4. Add 1 ml of PBS buffer to yeast pellet

Make Fermentation Solution

1. Mix the following ingredients for each lane