THE BEST METHOD WE HAVE FOUND (modify from Lisa Vaillancourt Lab)
(Method of John Tuite, modified by Rick Latin, 9-1-95)
1. Fill vials (cryovials Wheaton 8 ml, cat 224884 we get them from VWR) half full with silica gel granules, put caps on loosely, and sterilize by heating in 16o C drying oven (kitchen) for at least 2 hours. DO NOT AUTOCLAVE THE SILICA, IT WILL BECOME WET! (silica gel, Fisher, S156-212 , or silica gel desiccant 63231-67-4 Chemat http://www.chemat.com.pl/ will also work).
2. Prepare a 7.5% solution of Difco skim milk, autoclave. ( in Spain we use MERCK 1.15363.0500 skim milk powder)
3. Grow C. graminicola on Difco potato dextrose agar plates for 1-2 weeks under the lights. Do not use plates more than 2 weeks old.
4. Add 2 or 3 mls of sterile skim milk to the culture, GENTLY scrape the surface with a sterile loop to loosen the spores.
5. Add 0.5 mL of the milk-spore suspension to a vial of chilled, sterilized silica. Add the suspension slowly and evenly. Shake the vial in a vortex during and after the process of adding the suspension.
6. Tighten the cap on the vial, seal with parafilm. Label the vial!!! Use the small circular paper labels.
7. Vials can be stored at 4°C for a few months, or at -80°C for many years (up to 25 years and counting!). note: Need special cryo boxes for 8 ml vials.
8. To revive cultures, sprinkle a few silica grains onto a potato dextrose agar plate. The cultures may take a bit longer than normal to start growing.