# 169: TERETINEMA MEDIUM


## Main sol. 169

960	ml	**Solution A**	
20	ml	**Solution B**	
20	ml	**Solution C**	

1. Dissolve ingredients of solution A (except cysteine) and sparge medium with 100% N2 gas for 30 - 45 min to make it anoxic. Add cysteine and adjust pH 7.2 with KOH, then distribute under the same gas atmosphere in anoxic Hungate-type tubes or serum vials and autoclave. Solution B is autoclaved separately under 80% N2 and 20% CO2 gas atmosphere and solution C is autoclaved under 100% N2 gas atmosphere. To complete the medium appropriate amounts of solutions B and C are added to the sterile solution A in the sequence as indicated. Final pH of the complete medium should be 7.5.
2. For solid medium add 10.00 g/l agar (BD Bacto) to solution A.


## Solution A

0.04	g	CaCl2 x 2 H2O	
0.5	g	MgSO4 x 7 H2O	
4	g	Yeast extract (OXOID)	
2	g	D-Glucose	
0.5	ml	Sodium resazurin (0.1% w/v)	
0.5	g	L-Cysteine HCl x H2O	
10	g	Agar, for solid medium (optional)	
960	ml	Distilled water	


## Solution B

1	g	NaHCO3	
20	ml	Distilled water	


## Solution C

16.04	ml	K2HPO4 (0.5 M)	
3.96	ml	KH2PO4 (0.5 M)	

1. Combine 16.04 ml of 0.5 M K2HPO4 with 3.96 ml of 0.5 M KH2PO4 stock solution to reach a pH of around 7.4.