# 144a: CALDICELLULOSIRUPTOR ACETIGENUS MEDIUM


## Main sol. 144a

0.9	g	NH4Cl	
0.9	g	NaCl	
0.4	g	MgCl2 x 6 H2O	
0.75	g	KH2PO4	
1.5	g	K2HPO4	
9	ml	**Trace element solution**	
3	ml	FeSO4 x 7 H2O (0.1% w/v in 0.1 N H2SO4)	
1	g	Yeast extract	
0.5	ml	Sodium resazurin (0.1% w/v)	
5	ml	**Wolin's vitamin solution**	
5	g	D-Glucose	
1	g	Na2S x 9 H2O	
1000	ml	Distilled water	

1. Dissolve ingredients (except vitamins, glucose and sulfide), sparge medium with 100% N2 gas for 30 - 45 min to make it anoxic. Dispense medium under same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Add glucose, vitamins (sterilized by filtration) and sulfide from sterile anoxic stock solutions prepared under 100% N2 gas. Adjust pH of complete medium to 7.2 - 7.4.


## Trace element solution

12.8	g	Nitrilotriacetic acid (NTA)	
0.2	g	FeCl2 x 4 H2O	
0.1	g	MnCl2 x 4 H2O	
0.17	g	CoCl2 x 6 H2O	
0.1	g	CaCl2 x 2 H2O	
0.1	g	ZnCl2	
0.02	g	CuCl2	
0.01	g	H3BO3	
0.01	g	Na2MoO4 x 2 H2O	
0.03	g	NiCl2 x 6 H2O	
1	g	NaCl	
0.03	g	Na2SeO3 x 5 H2O	
1000	ml	Distilled water	

1. First dissolve NTA in 200 ml of distilled water and adjust pH to 6.5 with KOH, then dissolve mineral salts. Finally adjust pH to 6.5 with KOH and make up to 1000.00 ml.


## Wolin's vitamin solution

2	mg	Biotin	
2	mg	Folic acid	
10	mg	Pyridoxine hydrochloride	
5	mg	Thiamine HCl	
5	mg	Riboflavin	
5	mg	Nicotinic acid	
5	mg	Calcium D-(+)-pantothenate	
0.1	mg	Vitamin B12	
5	mg	p-Aminobenzoic acid	
5	mg	(DL)-alpha-Lipoic acid	
1000	ml	Distilled water