Solution A
Na2SO4……………………………………….......3.0 g
KH2PO4…………………………………………...0.2 g
NH4Cl……………………………………….…….0.3 g
NaCl………………………………………………1.0 g
MgCl2 x 6H2O…………………………………….0.4 g
KCl………………………………………………...0.5 g
CaCl2 x 2H2O……………………………………..0.15 g
DI Water…………………………………………...870.0 ml
Solution B
Trace elements SL-7 (see below)……………...1.0 ml
Solution C
NaHCO3……………………………………………5.0 g
DI Water……………………………………………100.0 ml
Solution D
Sodium propionate…………………………...........1.5 g
DI Water…………………………………………….10.0 ml
Solution E
Wolfe's Vitamin solution (see below)…………....10.0 ml
Solution F
Na2S x 9H2O……………………………………….0.4 g
DI Water……………………………………………10.0 ml
Solution A is prepared and autoclaved under 80%N2 + 20%CO2.Solutions B, D, E, and F are autoclaved separately; under nitrogen. Solution C is filter-sterilized and flushed with 80% N2 + 20%CO2 to remove dissolved oxygen.
Solutions B through F are added to sterile, cooled Solution A in the sequence indicated.
The complete medium is distributed anaerobically under 80% N2 + 20% CO2. Final pH of medium is 7.1 - 7.4.
Trace Element Solution SL-7
Hydrochloric acid, 25%.......................................10.0 ml
FeCl2 x 4H2O……………………………………..1.5 g
CoCl2 x 6H2O……………………………………..190.0 mg
MnCl2 x 4H20……………………………………..100.0 mg
ZnCl2………………………………………………70.0 mg
2821 H2BO4……………………………………………...62.0 mg
Na2MoO4 x 2H2O………………………………...36.0 mg
NiCl2 x 6H2O……………………………………....24.0 mg
CuCl2 x 2H2O……………………………………..17.0 mg
DI Water……………………………………………1000 ml
Dissolve FeCl2 in the hydrochloric acid, then dilute with water.
Wolfe's Vitamins
Folic acid……………………………………………2.0 mg
Pyridoxine hydrochloride…………………………10.0 mg
Riboflavin…………………………………………..5.0 mg
Biotin………………………………………………..2.0 mg
Thiamine…………………………………………...5.0 mg
Nicotinic acid………………………………………5.0 mg
Pantothenic acid…………………………………..5.0 mg
Vitamin B12………………………………………..0.1 mg
p-Aminobenzoic acid……………………………..5.0 mg
Thioctic acid………………………………………..5.0 mg
DI Water…………………………………………….1000 ml