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DSMZ

DSMZ 337 . SYNTROPHOCOCCUS SUCROMUTANS MEDIUM

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Equipment needed: Autoclave Centrifuge Filter Gassing station Hungate tubes
Final gas composition: 80 % N2 20 % CO2 read more
Final pH: 6.4 - 6.8
Select a strain (optional):

ml
Compound Amount Unit
Solution A 921 ml
Solution B 50 ml
Solution C 1 ml
Solution D 25 ml
Solution E 10 ml
Solution F 1 ml
1 Sparge solution A with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Adjust pH to 6.4 and dispense under same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Solution B is autoclaved separately under 80% N2 and 20% CO2 gas atmosphere. Solution C is prepared under 100% N2 gas atmosphere and sterilized by filtration. Solutions D, E and F are autoclaved separately under 100% N2 gas. To complete the medium add appropriate amounts of solutions B - F to the sterile solution A in the sequence as indicated. Adjust pH of the complete medium to 6.4 - 6.8, if necessary.
2 Note: Rumen fluid may be replaced by supplementing the medium with 200 µg/ml of crude egg yolk phosphatidylcholine (SIGMA, type IX-E).

Solution A #

921
Compound Amount Unit
Mineral solution 50.0 ml
Trace element solution SL-10 1.0 ml
Clarified rumen fluid 300.0 ml
Trypticase peptone
(BD Bacto/BD BBL) 		5.0 g
Na-formate 0.6 g
Sodium resazurin
(0.1% w/v) 		0.5 ml
Distilled water 570.0 ml

Solution B #

50
Compound Amount Unit
NaHCO3 2.5 g
Distilled water 50.0 ml

Solution C #

1
Compound Amount Unit
Seven vitamins solution 1 ml

Solution D #

25
Compound Amount Unit
Lactose 5 g
Distilled water 25 ml

Solution E #

10
Compound Amount Unit
L-Cysteine HCl x H2O 0.24 g
Distilled water 10.00 ml

Solution F #

1
Compound Amount Unit
Na2S x 9 H2O 78 mg
Distilled water 1 ml

Mineral solution #

1000
Compound Amount Unit
KH2PO4 10.0 g
MgCl2 x 6 H2O 6.6 g
NaCl 8.0 g
NH4Cl 8.0 g
CaCl2 x 2 H2O 1.0 g
Distilled water 1000.0 ml

Trace element solution SL-10 #

1000
Compound Amount Unit
HCl
(25%) 		10.0 ml
FeCl2 x 4 H2O 1.5 g
ZnCl2 70.0 mg
MnCl2 x 4 H2O 100.0 mg
H3BO3 6.0 mg
CoCl2 x 6 H2O 190.0 mg
CuCl2 x 2 H2O 2.0 mg
NiCl2 x 6 H2O 24.0 mg
Na2MoO4 x 2 H2O 36.0 mg
Distilled water 990.0 ml
1 First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally make up to 1000.00 ml.

Clarified rumen fluid #

1000
1 Rumen fluid from cow or sheep (obtained from fistulated animals or abattoir refuse) is filtered through muslin, autoclaved at 121°C for 15 min and then centrifuged at 27,000 g for 20 min. The supernatant is made anoxic by sparging with 100% N2 gas for 15 min, dispensed under same gas atmosphere into anoxic serum vials to 30% of volume and then stored frozen at -20°C.

Seven vitamins solution #

1000
Compound Amount Unit
Vitamin B12 100 mg
p-Aminobenzoic acid 80 mg
D-(+)-biotin 20 mg
Nicotinic acid 200 mg
Calcium pantothenate 100 mg
Pyridoxine hydrochloride 300 mg
Thiamine-HCl x 2 H2O 200 mg
Distilled water 1000 ml