Rehydration and Growth of freeze dried cultures
Each yeast is dispatched as a small quantity of freeze-dried culture sealed under vacuum in glass ampoules. Ampoules should be opened in the following way:
- Check the number on the label inside the ampoule.
- Score the glass with a suitable file or scoring impliment at the level of the cotton wool plug.
- Wipe the ampoule with alcohol soaked tissue, then wrap the tissue around the ampoule.
- Using sufficient padding, apply pressure both sides of the mark and snap the ampoule along the score line. Be careful in this step to avoid a glass stab wound.
- Remove the tip of the ampoule and cotton wool plug and place in a sharp container.
- Using a sterile Pasteur pipette, add about 0.5ml YM broth (Difco 0711-01) or malt extract to the dried material.
- Gently resuspend the dried culture and transfer to a culture bottle containing approximately 10ml of the same medium.
- Unless otherwise indicated, most yeasts should be incubated at 25°C. Growth may be slow immediately after resuscitation. It may be necessary to incubate cultures for at least 5 days before discarding. Growth is usually stimulated by aeration which may be achieved by shaking or rocking the culture.
- The discarded ampoule should be placed in a container for subsequent sterilization before disposal.
All NCYC strains (except where indicated) can be routinely grown in YM broth (Difco 0711-01). Other suitable nutrient media include:
- YM: 0.3% yeast extract, 0.3% malt extract, 0.5% peptone, 1% glucose
- YEP-glucose: 0.5% yeast extract, 0.5% peptone, 1% glucose
- Malt Extract: 0.3% malt extract, 0.5% peptone
- Sabouraud’s Glucose: 1% peptone, 4% glucose
- YPD medium: 1% yeast extract, 2% peptone, 2% glucose
- Yeast Nitrogen Base (Difco 0392-15-9): a chemically defined medium to which a carbon source must be added.
- Yeast Nitrogen Base without amino acids (Difco 0919-15-3), which can be supplemented with appropriate amino acids or other source of nitrogen (useful for some genetically defined strains). A carbon source must be added.
Agar when required is added to a final concentration of 1.5 – 2.0% (All percentages are given weight/volume)
More information about these media is given by J P van der Walt & D Yarrow, ‘Methods for the isolation, maintenance, classification and identification of yeasts’ in ‘The Yeasts . A Taxonomic Study’, third edition by ed. N J W Kreger-van Rij, Elsevier, (1984) pp 45-104, and in ‘Yeasts: Characteristics and Identification’, third edition by J A Barnett, R W Payne & D Yarrow, Cambridge University Press, (2000).