# 9a: VY/2, REDUCED MEDIUM


## Main sol. 9a

2.5	g	Baker's yeast	
1.36	g	CaCl2 x 2 H2O	
0.5	mg	Vitamin B12	
8	g	Agar (Difco)	
1000	ml	Distilled water	

1. Sterilize vitamin B12 separately by filtration. Prepare and store yeast cells as autoclaved stock suspension (5 g baker's yeast/100 ml distilled water, adjust pH to 6.5 and autoclave).
2. Adjust pH of medium to 7.2 with KOH before as well as after autoclaving and cooling to 50°C (use pH-indicator paper).
3. For suspension of freeze-dried cells from ampoules add about 0.5 - 1.0 ml medium MD1 (per liter: casiton, 3.0 g; CaCl2 x 2 H2O,  0.7 g; MgSO4 x 7 H2O, 2.0 g) to the vial with freeze dried material.