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Main sol. 1596

Internal Solution-ID: S3295

Compound	Amount	Unit	Conc. [g/L]	Conc. [mM]
NaCl	3.00	g	2.991	51.181
K₂HPO₄	1.00	g	0.997	5.724
NaHCO₃	20.00	g	19.94	237.363
Na₂CO₃	15.00	g	14.955	141.102
NH₄Cl	0.20	g	0.199	3.728
MgSO₄ x 7 H₂O	0.50	g	0.499	2.023
Trace elements solution (Pfennig & Lippert,1966)	1.00	ml	-	-
Selenite-tungstate solution	1.00	ml	-	-
Na-formate	3.40	g	3.39	49.845
Na-acetate	0.16	g	0.16	1.945
Yeast extract (OXOID)	0.02	g	0.02	-
Wolin's vitamin solution (10x)	1.00	ml	-	-
Na₂S x 9 H₂O	0.25	g	0.249	1.038
Distilled water	1000.00	ml	-	-
1 Dissolve sodium chloride and hydrogenphosphate, then sparge solution with 100% N₂ gas for at least 30 - 45 min to make it anoxic. Add and dissolve hydrogencarbonate and carbonate while gassing the head space only, then dispense under 100% N₂ gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. After autoclaving add ammonium chloride, magnesium sulfate, trace elements, formate, acetate, yeast extract, vitamins and sulfide from sterile anoxic stock solutions prepared under 100% N₂ gas. Vitamins should be sterilized by filtration. The pH of the complete medium should be 9.5.
2 Note: Addition of 10 - 20 mg sodium dithionite per liter (e.g. from 5% (w/v) solution, freshly prepared under N₂ and filter-sterilized) may stimulate growth at the beginning. For transfers use 5 - 10% inoculum.

Compound

Amount

Unit

Conc. [g/L]

Conc. [mM]

NaCl

3.00

2.991

51.181

K₂HPO₄

1.00

0.997

5.724

NaHCO₃

20.00

19.94

237.363

Na₂CO₃

15.00

14.955

141.102

NH₄Cl

0.20

0.199

3.728

MgSO₄ x 7 H₂O

0.50

0.499

2.023

Trace elements solution (Pfennig & Lippert,1966)

1.00

Selenite-tungstate solution

1.00

Na-formate

3.40

3.39

49.845

Na-acetate

0.16

1.945

Yeast extract
(OXOID)

0.02

Wolin's vitamin solution (10x)

1.00

Na₂S x 9 H₂O

0.25

0.249

1.038

Distilled water

1000.00

1 Dissolve sodium chloride and hydrogenphosphate, then sparge solution with 100% N₂ gas for at least 30 - 45 min to make it anoxic. Add and dissolve hydrogencarbonate and carbonate while gassing the head space only, then dispense under 100% N₂ gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. After autoclaving add ammonium chloride, magnesium sulfate, trace elements, formate, acetate, yeast extract, vitamins and sulfide from sterile anoxic stock solutions prepared under 100% N₂ gas. Vitamins should be sterilized by filtration. The pH of the complete medium should be 9.5.

2 Note: Addition of 10 - 20 mg sodium dithionite per liter (e.g. from 5% (w/v) solution, freshly prepared under N₂ and filter-sterilized) may stimulate growth at the beginning. For transfers use 5 - 10% inoculum.

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Main sol. 1596

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