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• Homogenization of Candida albicans for Protein Extraction

Homogenization of Candida albicans for Protein Extraction

Instrument: Bullet Blender or Bullet Blender Blue

Sample: C. albicans culture

Protocol:

1. Take 50ml of C. albicans culture at an OD600nm of 1.0.
2. Harvest by vacuum filtration, resuspend in ice-cold 20% TCA, and incubate on ice for 30min.
3. Centrifuge culture to yield a cell pellet.
4. Wash the pellet 2x in alkaline-buffered acetone (three parts 3M Tris, pH 8.8, to seven parts acetone).
5. Air-dry the pellet. Resuspend in 8M urea.
6. Add a volume of 0.5mm zirconium oxide beads equal to the volume of the pellet.
7. Run the samples in the Bullet Blender at speed 8 for 5 minutes.
8. Remove tubes from the instrument and visually inspect the samples. If homogenization is unsatisfactory, run for another two minutes at speed 10.

Reference:  This protocol is adapted from: Rauceo JM, Blankenship JR, Fanning S, Hamaker JJ, Deneault JS, Smith FJ, Nantel A, Mitchell AP. Regulation of the Candida albicans cell wall damage response by transcription factor Sko1 and PAS kinase Psk1. Mol Biol Cell. 2008 Jul;19(7):2741-51.