UW Stout/Ethanol FA19

Yeast torture 1

Protocol:

Materials:

1. neat ethanol
2. yeast (WT:BY4735)
3. sterile water
4. 200 ul
5. 96 well plate

Procedure:

1. (add 50ul mixture to 50ul yeast in 96 well plate.)x2. Used 12 wells in total
2. fill 6 200ul tubes, each with water and ethanol concentration x2. Resulting in 6 200ul tubes with 100ul of mixture in each.

	1	2	3	4	5	6
ul ethanol	0	3.75	7	10.25	13.5	16.75
ul water	50	46.25	43	39.75	37.5	33.25
ul yeast	50	50	50	50	50	50

This graph shows the ethanol tolerance in the Wild Type Yeast Cells (BY4735). This graph helped us determine the amount of yeast to add to our knockout strains.

Knockout Strand Yeast Torture

Protocol:

Materials:

1. Yeast Knockout Strains:
   1. YAL061W
   2. YBL086C
   3. YBL9081W
   4. YBR033W
   5. YBR138C
   6. YBR220C
   7. YHR048W
   8. YGL140C
   9. YJR061W
   10. YKR005C
   11. WT:BY4735
2. neat ethanol
3. sterile water
4. micropipeter(s)
5. 96 well plate

Procedure:

1. add 50 ul of Knockout Strain to individual well (96 well plate).
2. add 50 ul of water+ethanol mixture to individual well (Follow table below).

Well	1	2	3	4	5	6	7	8	9	10	11	12
ul yeast	50	50	50	50	50	50	50	50	50	50	50	50
ul ethanol	7	7	7	7	7	7	7	7	7	7	7	7
ul water	43	43	43	43	43	43	43	43	43	43	43	43