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Solution A

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Internal Solution-ID: S4692

Equipment needed: Anaerobic workbench Filter Autoclave
Compound Amount Unit Conc. [g/L] Conc. [mM]
Na2SO4 0.700 g 0.684 4.813
K2HPO4 0.200 g 0.195 1.121
NH4Cl 0.250 g 0.244 4.564
NaCl 0.250 g 0.244 4.178
MgCl2 x 6 H2O 0.400 g 0.391 1.921
KCl 0.500 g 0.488 6.55
CaCl2 x 2 H2O 0.150 g 0.146 0.996
Resazurin 0.500 mg 4.9e-4 0.002
Yeast extract 2.000 g 1.953 -
FeCl2 solution 1.000 ml - -
Trace element solution 1.000 ml - -
Selenite-tungstate solution 1.000 ml - -
L-Cysteine HCl x H2O 0.050 g 0.049 -
FeSO4 x 7 H2O 0.025 g 0.024 0.088
Distilled water 900.000 ml - -
1 Mix components except L-cysteine x HCl x H2O and FeSO4 x 7H2O. Bring to a boil for a few seconds and cool down under a N2 atmosphere. While gassing with N2, add L-cysteine x HCl x H2O and FeSO4 x 7H2O to the solution, distribute into culture vessels (e.g., 18 ml in 50 ml serum bottles) under a N2 atmosphere, then, seal butyl rubber stoppers and autoclave. Aseptically and anaerobically add per liter (final) the following sterile solutions (autoclaved or *filter-sterilized) from anaerobic stocks:
Potassium phosphate buffer , pH 7.5
(0.1 M) 		10.000 ml - -
Trace vitamins (see Medium No. 197)
(*) 		10.000 ml - -
Vitamin B12 solution (see Medium No. 403)
(*) 		1.000 ml - -
NaHCO3
(*, 8%) 		42.000 ml - -
Sodium pyruvate
(*, 25%) 		18.000 ml - -
Sodium fumarate
(*, 16%) 		40.000 ml - -
2 Adjust pH of the medium to 7.3 - 7.7, if necessary.
J753 DEHALOSPIRILLUM MEDIUM