Difference between revisions of "UW-Stout/Caffeine FA22"
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1. Vortex the yeast briefly before use. | 1. Vortex the yeast briefly before use. | ||
| − | 2. Add 50ul of the 0.0125 M stock solution and 50ul of wild type yeast to | + | 2. Add 50ul of the 0.0125 M stock solution and 50ul of wild type yeast to the first well. |
3. Repeat for each knockout gene in the subsequent wells. | 3. Repeat for each knockout gene in the subsequent wells. | ||
Revision as of 13:30, 29 November 2022
Contents
Yeast Caffeine Torture
Caffeine Solubility: 2g/ 100 mL
Pilot Experiment Protocol
Materials:
- Sterile Water
- Caffeine
- Wild Yeast
- Weigh Boat
- Microcentrifuge tubes
- Ethanol spray to sterilize equipment
Procedure:
Stock Solutions
Underneath the hood...
1. Make a stock solution at a concentration of 0.1 M
- Add 0.19419 g of caffeine to 10 mL water
- Vortex briefly
2. Make a stock solution at a concentration of 0.05 M
- Take 250 ul of 0.1 M and add 250 ul water
- Vortex briefly
3. Make a stock solution at a concentration of 0.025 M
- Take 250 ul of 0.05 M and add 250ul of water
- Vortex briefly
4. Make a stock solution at a concentration of 0.0125 M
- Take 250 ul of 0.025 M and add 250 ul water
- Vortex briefly
5. Make a stock solution at a concentration of 0.00625 M
- Take 250 ul of 0.0125 and add 250 ul of water
- Vortex briefly
6. Make a stock solution at a concentration of 0.003125 M
- Take 250 ul of 0.00625 M and add 250 ul of water
- Vortex briefly
7. Make a stock solution at a concentration of 0.0015625 M
- Take 250 ul of 0.003125 M and add 250 ul of water
- Vortex briefly
8. Make a stock solution at a concentration of 7.8125 x 10^-4 M
- Take 250 ul of 0.0015625 M and add 250 ul of water
- Vortex briefly
9. Make a stock solution at a concentration of 3.90625 x 10^-4 M
- Take 250 ul of 7.8125 x 10^-4 M and add 250 ul of water
10. Make a stock solution at a concentration of 1.953125 x 10^-4 M
- Take 250 ul of 3.90625 x 10^-4 ul and add 250 ul of water
- Vortex briefly
Yeast + Wells
- Vortex the yeast briefly before use.
- To the first well, add 50 ul of 0.1 M stock solution and 50 ul of yeast.
- Repeat the first step in the next well for 0.05 M solution, continuing to add each stock solution to the next wells.
Inducing Stress Protocol
Materials
- 10 mL 0.1 M Stock Solution
- Sterile Water
- Test tubes
- Yeast sample
Procedure
Stock Solution
1. Make 1 ml of stock solution at a concentration of 0.0125 M
- Add 125ul of the 10 ml 0.1M stock solution to 875ul of sterile water
- Vortex Briefly
Yeast + Wells
1. Vortex the yeast briefly before use.
2. Add 50ul of the 0.0125 M stock solution and 50ul of wild type yeast to the first well.
3. Repeat for each knockout gene in the subsequent wells.
