UW-Stout/Caffeine SP21

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Materials

  • Caffeine 80 mM solution in water. (15.52 x 25 ml water= .388 g of caffeine)
  • Sterile H2O
  • Wild-type yeast in 2x synthetic complete media at an OD600 of 0.1-0.2
  • Corning COSTAR 96-well clear flat-bottom assay plate
  • 250 mL beaker for caffeine master solution.
  • P-10,200 micro pipette/ tips
  • Scale
  • Tube to store caffeine
  • Powdered caffeine for solution


Equiptment

  • Molecular Devices SpectraMax Plus 384 Microplate Reader


Protocol

  1. Weigh out .388 grams of caffeine
  2. Add to 25 ml of water in test tube and swirl until it’s mixed with no chunks
  3. Set up 8 test tubes and label them 1 through 8 matching with the order of wells
  4. Using the p20 pipette, pipette the varying amounts of caffeine solution provided in table
  5. Then using a p200 pipette, pipette the varying amount of sterile water into the provided tubes
  6. When under the hood add 50 ul of yeast to each test tube except for the tube labeled 0 which is the control
  7. Transfer the solution created in each tube into the wells, writing down which row your wells are
  8. Set up the plate reader as follows:

Temperature: 30°C Mode: Kinetic Wavelength: 600 nm Interval: 10 minutes Total run time: 12 hours Shake before read: 30 seconds Transfer the assay plate to the reader and read for 24 hours. This process will be replicated 3 times.

Data

[[File:]]