Materials and Reagents

1. Serapias vomeracea seeds (collected in the field, in the north-west Mediterranean meadows of Italy; seeds from this orchid species could be retrieved through “index seminum” of international botanic gardens or germplasm banks)
   
2. Tulasnella calospora inoculum (strain MUT 4182) (previously grown in sterile culture on MEA medium)
   
3. Sterile double deionized water (neutral pH)
   
4. Tween-20 (Sigma-Aldrich, catalog number: P2287 )
   
5. Milled oats (retrieved from organic shops)
   
6. Malt Extract (Sigma-Aldrich, Fluka, catalog number: 70167 )
   
7. Ethanol (Sigma-Aldrich, Fluka, catalog number: 0 2860 )
   
8. 5% sodium hypochlorite (commercial bleach)
   
9. Agar (Sigma-Aldrich, catalog number: A1296 )
   
10. Glucose (Sigma-Aldrich, catalog number: G8270 )
    
11. Peptone (Sigma-Aldrich, Fluka, catalog number: P5905 )
    
12. Sterilizing solution (see Recipes)
    
13. Solid oat medium (see Recipes)
    
14. MEA medium (see Recipes)
    

Equipment

1. Balance
   
2. Small spatula
   
3. Small metal forceps
   
4. Sterile 10 x 10 mm Whatman No.1 filter paper
   
5. Lancets
   
6. 2.0 ml centrifuge tubes
   
7. Micropipette
   
8. Filter tips
   
9. Centrifuge
   
10. Vortex
    
11. Timer
    
12. Laminar flow hood
    
13. Petri dishes (9 cm diameter)
    
14. Parafilm (Sigma-Aldrich, catalog number: P7793 )
    
15. Bunsen burner
    
16. Climatic chamber (Binder GmbH)
    

Procedure