# 930: NATRONINCOLA HISTIDINOVORANS MEDIUM


## Main sol. 930

0.2	g	KH2PO4	
0.1	g	MgCl2 x 6 H2O	
1	g	NH4Cl	
0.2	g	KCl	
80	g	NaCl	
6.83	g	Na2CO3	
3.83	g	NaHCO3	
0.2	g	Na2S x 9 H2O	
0.2	g	Yeast extract	
5	g	Histidine	
2	ml	**Wolin's vitamin solution**	
1	ml	**Modified Wolin's mineral solution II**	
1	mg	Resazurin	
1000	ml	Distilled water	

1. Final pH 8.9
2. Prepare the medium without adding the vitamins, histidine, yeast extract, NH4Cl, Na2CO3, and NaHCO3 using anaerobic conditions, under nitrogen. If the medium has been boiled to remove oxygen add the NH4Cl, Na2CO3, and NaHCO3 after the medium had cooled. Dispense into tubes stopper with rubber stoppers (serum tubes or bottles, or screw capped tubes). Autoclave, and to the cooled medium add the vitamins (filter sterilised under N2), histidine, yeast extract, and Na2S x 9 H2O from anaerobic, sterile stock solutions.


## 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	


## Modified Wolin's mineral solution II

1.5	g	Nitrilotriacetic acid	
3	g	MgSO4 x 7 H2O	
0.5	g	MnSO4 x H2O	
1	g	NaCl	
0.1	g	FeSO4 x 7 H2O	
0.18	g	CoSO4 x 7 H2O	
0.1	g	CaCl2 x 2 H2O	
0.18	g	ZnSO4 x 7 H2O	
0.01	g	CuSO4 x 5 H2O	
0.02	g	AlK(SO4)2 x 12 H2O	
0.01	g	H3BO3	
0.01	g	Na2MoO4 x 2 H2O	
0.025	g	NiCl2 x 6 H2O	
0.3	mg	Na2SeO3 x 5 H2O	
1000	ml	Distilled water	

1. First dissolve nitrilotriacetic acid and adjust pH to 6.5 with KOH, then add minerals. Adjust final to pH 7.0 with KOH.