Difference between revisions of "UW-Stout/Hydroxychloroquine SP21"
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==Equipment== | ==Equipment== | ||
* [https://www.moleculardevices.com/sites/default/files/en/assets/data-sheets/br/spectramax-plus-384-microplate-reader.pdf Molecular Devices SpectraMax Plus 384 Microplate Reader] | * [https://www.moleculardevices.com/sites/default/files/en/assets/data-sheets/br/spectramax-plus-384-microplate-reader.pdf Molecular Devices SpectraMax Plus 384 Microplate Reader] | ||
| + | |||
| + | ==Protocol== | ||
| + | 1. Vortex the yeast culture briefly to resuspend the yeast cells. | ||
| + | 2. Set up eight wells according to the following table: | ||
| + | |||
| + | [[File:Capture.png]] | ||
| + | |||
| + | The final concentration of hydroxychloroquine is 300ug/mL. We have controls for this experiment: a well with the wild-type strain yeast untreated with the hydroxychloroquine solution, and wells with the modified strains yeast untreated with the hydroxychloroquine solution for all 8 modified strains. We also have experimental samples: a well with the wild-type strain yeast treated with the hydroxychloroquine solution, and wells with the modified strains yeast treated with the hydroxychloroquine solution for all 8 modified strains. | ||
| + | 3. To measure cell growth over a period of 24 hours, set up the plate reader as follows: | ||
| + | Temperature: 30°C | ||
| + | Mode: Kinetic | ||
| + | Wavelength: 600 nm | ||
| + | Interval: 5 minutes | ||
| + | Total run time: 24 hours | ||
| + | Shake before reading: 30 seconds | ||
| + | 4. Transfer the assay plate to the reader and read for 24 hours. | ||
| + | Note: Repeat the experiment a few times to account for natural variation. | ||
Revision as of 08:02, 27 April 2021
Materials
- Corning COSTAR 96-well clear flat-bottom assay plate
- Wild-type yeast in 2x synthetic complete media at an OD600 of 0.1-0.2
- Hydroxychloroquine, 300 ug/uL solution in sterile water
- P20, P200 micropipettors
Equipment
Protocol
1. Vortex the yeast culture briefly to resuspend the yeast cells. 2. Set up eight wells according to the following table:
The final concentration of hydroxychloroquine is 300ug/mL. We have controls for this experiment: a well with the wild-type strain yeast untreated with the hydroxychloroquine solution, and wells with the modified strains yeast untreated with the hydroxychloroquine solution for all 8 modified strains. We also have experimental samples: a well with the wild-type strain yeast treated with the hydroxychloroquine solution, and wells with the modified strains yeast treated with the hydroxychloroquine solution for all 8 modified strains. 3. To measure cell growth over a period of 24 hours, set up the plate reader as follows: Temperature: 30°C Mode: Kinetic Wavelength: 600 nm Interval: 5 minutes Total run time: 24 hours Shake before reading: 30 seconds 4. Transfer the assay plate to the reader and read for 24 hours. Note: Repeat the experiment a few times to account for natural variation.
