ATCC medium: 1937 Modified DSM medium 503 with xylan
Solution A:
KH2PO4 ...................................0.2 g
NH4Cl ....................................0.25 g
NaCl.....................................1.0 g
MgCl2 . 6H2O .............................0.4 g
KCl......................................0.5 g
CaCl2 . 2H2O .............................0.15 g
Xylan....................................2.0 g
Modified Wolfe's Minerals (see below)...10.0 ml
Resazurin................................0.5 mg
Distilled water........................940.0 ml
Prepare basal medium anaerobically under 80% N2, 20% CO2. Aseptically
add the following sterile stock solutions to the cooled, sterile basal
medium. Adjust medium for final pH 7.3 +/- 0.1.
Solution B:
Wolfe's Vitamin Solution (see below)....10.0 ml
Autoclave for 15 minutes at 121C under 100% N2.
Solution C:
5% NaHCO3 ...................50.0 ml
Autoclave for 15 minutes at 121C under 100% N2.
Solution D:
Na2S . 9H2O ..................0.3 g
Distilled water.............10.0 ml
Autoclave for 15 minutes at 121C under 100% N2.
Modified Wolfe's Minerals:
Na2SeO3 .................................10.0 mg
NiCl2 . 6H2O ............................10.0 mg
Na2WO4 . 2H2O............................10.0 mg
Wolfe's Mineral Solution (see below).....1.0 L
Wolfe's Mineral Solution:
Available from ATCC as a sterile ready-to-use liquid (Trace Mineral
Supplement, catalog no. MD-TMS.)
Nitrilotriacetic acid........1.5 g
MgSO4 . 7H2O .................3.0 g
MnSO4 . H2O ..................0.5 g
NaCl.........................1.0 g
FeSO4 . 7H2O .................0.1 g
CoCl2 . 6H2O .................0.1 g
CaCl2 ........................0.1 g
ZnSO4 . 7H2O .................0.1 g
CuSO4 . 5H2O .................0.01 g
AlK(SO4)2 . 12H2O.............0.01 g
H3BO3 ........................0.01 g
Na2MoO4 . 2H2O................0.01 g
Distilled water..............1.0 L
Add nitrilotriacetic acid to approximately 500 ml of water and adjust to
pH 6.5 with KOH to dissolve the compound. Bring volume to 1.0 L with
remaining water and add remaining compounds one at a time.
Wolfe's Vitamin Solution:
Available from ATCC as a sterile ready-to-use liquid (Vitamin
Supplement, catalog no. MD-VS).
Biotin.......................2.0 mg
Folic acid...................2.0 mg
Pyridoxine hydrochloride....10.0 mg
Thiamine . HCl...............5.0 mg
Riboflavin...................5.0 mg
Nicotinic acid...............5.0 mg
Calcium D-(+)-pantothenate...5.0 mg
Vitamin B12..................0.1 mg
p-Aminobenzoic acid..........5.0 mg
Thioctic acid................5.0 mg
Distilled water..............1.0 L