In this article we will discuss about solid media preparation (PDA) for obtaining fungal culture in the pure form
Fungi are heterotrophic in nature which means that they are not capable of photosynthesis and thus require organic matter for growth and energy formation. Fungi can live as saprophytes on dead plants and animals or their wastes or parasites assimilating tissues of living plants and animals. A typical fungal life cycle features formation of threadlike vegetative hyphae which form a mycelium, a three-dimensional structure of hyphae capable of effective assimilation of nutrients and aggressive growth. Hyphae emerge from germinating spores (conidia) that may be uni- or multinucleate, haploid or diploid.
Mushrooms are typically isolated by plating a sample (spores, tissues) on a Petri dish containing a rich medium such as malt extract agar and potato dextrose agar (PDA) supporting the growth of a variety of mushrooms.
Needed tools and equipment for solid media preparation
1. 250 mL Erlenmeyer flasks and suitable caps. Cotton caps can be used, then covered by aluminum foil.
2. Sterile plastic or glass Petri dishes, vented. Standard size for fungal cultures is 9 cm in diameter and 1.3 cm or 2.0 cm in height.
3. Glass tube or sterile plastic tubes.
4. Graded glass containers; beakers for efficient mixing of the contents.
5. Small glass funnels plugged with permeable cotton wool, wrapped in foil and autoclaved using the dry program option.
6. Glass pipettes (10 mL) and measuring cylinders.
7. Autoclave (bench top, free standing, industrial, etc.) or Pressure cooker (optional).
8. Incubator cabinets set at 22-26 °C to grow the plate cultures.
9. Laminar hood.
Prepare PDA solid media
Components: Agar 20g; Potatoes (sliced or peeled) 200g; Dextrose 20g; Distilled water:1000ml.
Potato slices are allowed to simmer in 500 ml. distilled water, the extract filtered by means of a muslin cloth, dextrose and agar-agar powder are added, the volume made up to 1000 ml and made to boil.
Sterilization of the Medium (Autoclave): The media thus dispensed with should be steam-sterilized in an autoclave for 15 minutes at 121oC. For convenience, the culture tubes are arranged in a wire basket and then put in the autoclave. Before opening the autoclave, the pressure is allowed to come down to zero.
PDA petri plates
Cool the agar after autoclaving to about 60–70 °C for pouring of plates. 20ml of media was poured into petri dishes under laminar hood.
The culture tubes with 10-15 ml of medium and meant for use as tube slants should be taken out from the autoclave while hot, slightly rotated, put on a slanting position on a slanting rack and allowed to cool.
The medium due to the presence of agar-agar would solidify on cooling. The agar slants as well as plates are now ready for culture work.