UW-Stout/Prednisone FA19

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Revision as of 14:40, 10 December 2019 by Teagueb (talk | contribs) (Protocol)
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Materials

  • Prednisone
  • Absolute Ethanol
  • Yeast Cultures
  • Sterile Water
  • Corning COSTAR 96-Well Clear Flat-Bottom Assay Plate
  • Micropipettes
  • Centrifuge Tubes

Equipment

  • Incubator set to 30°C
  • Molecular Devices SpectraMax Plus 384 Microplate Reader

Protocol

  1. Make stock solution by vortexing the following:
    1. 50 mg Prednisone
    2. 10 ml ethanol
  2. Dilute down to needed percentages (below) using sterile water.
  3. In each well, add 50 µl of wildtype yeast strain.
  4. Using the stock solution (P1), add 20 µl to the first well along with 30 µl of sterile H2O.
    1. This well should now have 100 µl of total solution.
  5. Using the stock solution (P1), add 6 µl to the second well along with 44 µl of sterile H2O.
    1. This well should now have 100 µl of total solution.
  6. Repeat this process for each set of wells, going down one dilution after each set of two wells.
    1. Refer to the Well Concentrations chart.
  7. Dilute pure ethanol down to needed percentages (below).
  8. In 12 different wells, a new row, add 50 µl of wildtype yeast strain.
  9. Add the ethanol dilutions to the 12 new wells, using the same process as used above.
    1. This row will be used to compare the Prednisone treated yeasts’ growth rates.
  10. Shake all wells at 37°C for 24 hours.
  11. Plot data and select the Prednisone amount/concentration for the next test that shows results but doesn’t completely kill the yeast cultures.