Chlamydomonas meslinii (cells)
For the cryopreservation of microalgae. Protocol has been adapted from Muller, et al. (2007).
• Dimethylsulfoxide (DMSO)
• Controlled-rate cooler
• Cryostorage vessel with liquid nitrogen
• Waterbath at 45°C
1. Resuspend algal cultures in appropriate medium and adjust density to 1-5x107 cells/ml.
2. In a 2ml cryopreservation vial add 0.9ml cell culture with 10% DMSO (for a final concentration of 5% v/v DMSO)
3. Allow culture to rest for 10mins on ice, then transfer vials to a controlled-rate cooler, pre-chilled to 4°C.
4. Chill cultures at a rate of -1°C/min to a holding temperature of -35°C. Hold at -35°C for 40mins.
5. Again using a controlled cooling rate of -1°C/min, chill cultures to -45°C.
6. Transfer to a cryostorage vessel containing liquid nitrogen.
7. To thaw, submerge the vials in a 45°C water bath until culture is completely thawed (approximately 2-3mins).
8. Immediately dilute cultures 20-fold in fresh medium to reduce concentration of cryoprotective agent.
9. Culture algae following standard culturing procedures.
Muller, J., J.G. Day, K. Harding, D. Hepperle, M. Lorenz, and T. Friedl. 2007. Assessing genetic stability of a range of terrestrial microalgae after cryopreservation using amplified fragment length polymorphism (AFLP). Am J Bot. 94:799-808.
Added by: Carlton Hoyt on July 25, 2012
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