Preparation of Culture Media
Culture media are essential for isolation of mushroom fungi and maintaining them in a pure culture either in test tubes or in Petri plates. The fungal cultures may be grown in liquid media (broths) or on solid agar media. ( Agar agar is a solidifying material obtained from sea seed weed like Geladium sp., which melts on boiling and solidifies upon cooling).
Some of the commonly used media are listed below:
i) Potato dextrose agar (PDA):
| Potato |
250 g |
| Dextrose |
20 g |
| Agar agar |
20 g |
| Distilled water |
1000 ml. (pH : 7) |
ii) Oatmeal agar (OA)
| Oatmeal |
30 g |
| Agar agar |
20 g |
| Distilled water |
1000 ml. (pH : 7) |
iii) Wheat meal agar (WMA)
| Wheat grain |
30 g |
| Agar agar |
30 g |
| Distilled water |
1000 ml. (pH : 7) |
iv) Malt extract agar (MEA)
| Malt extract |
25 g |
| Agar agar |
20 g |
| Distilled water |
1000 ml. (pH : 7) |
v) Yeast potato dextrose agar (YPDA)
| Yeast granules |
1 g |
| Potato |
200 g |
| Dextrose |
20 g |
| Agar agar |
20 g |
| Distilled water |
1000ml (pH: 7) |
Potato dextrose agar (PDA):
Potato dextrose agar medium is very commonly used for isolation of fungus as well as for its maintenance.
Procedure:
- Wash 250 g potato, peel off the skin, and slice them into small pieces.
- Cook the sliced potato in 500 ml. water for 30 minutes in an open vessel or pressure cooker for 20 minutes.
- Simultaneously mix 20 g agar with 500 ml. of water and boil in a cooker for 30 minutes.
- Collect the potato extract by filter through muslin cloth or net filter.
- Add 20 g dextrose to the potato extract
- Mix thoroughly the molten agar with the potato- agar mixture and make the volume
to 1 litre with distilled water.
- Check the pH of the medium using pH papers.
- Pour the medium in to cleaned boiling tubes @ 15 ml / tube and plug with non-
absorbent cotton wool.
- Arrange the tubes in a wire basket, cover it with a waste paper sheet and tie tightly
with a cotton thread.
- Sterilize them in an autoclave or a pressure cooker at 15 lbs pressure for 20
minutes.
- Take out the sterilized tubes after releasing the steam and keep them in a slanting
position to get agar slants.
- After solidifying, the tubes are arranged in a wire basket and stored in a clean
room for further use.
Procedure for checking pH: Stir the medium with a glass rod and take a small bit of pH indicator paper and dip it in the medium. Take it out and compare the colour development with the colour chart given on the cover sheet of the filter paper book. If pH is above 7, add a few drops of 0.1N Hydrochloric acid (HCl), stir well and again check the pH for neutral. Contrastingly, if the pH of the medium is below 7, add a few drops of 0.1 N Sodium hydroxide (NaOH) to increase the pH to 7.
 |
| Boiling potatoes |
Boiling agar |
Potato extract + agar+ sugar |
 |
| Sterilization |
PDA media |
Pouring in Petri plates |
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