Marine Thermococcus Medium (DSMZ Medium 760)

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Marine Thermococcus Medium
(DSMZ Medium 760)

山东拓普生物工程有限公司 培养基配方 http://www.topbiol.com

Composition per liter:

NaCl..........................................................................................19.45g
MgCl 2 ............................................................................................8.8g
Sulfur............................................................................................5.0g
Peptone .........................................................................................5.0g
Na 2 SO 3 ........................................................................................3.24g
CaCl 2 .............................................................................................1.8g
Yeast extract..................................................................................1.0g
KCl..............................................................................................0.55g
NaHCO 3 ......................................................................................0.16g
Ferric citrate..................................................................................0.1g
KBr..............................................................................................0.08g
SrCl 2 ............................................................................................0.03g
H 3 BO 3 .........................................................................................0.02g
Na 2 HPO 4 ....................................................................................8.0mg
Na 2 SiO 3 ......................................................................................4.0mg
NaF.............................................................................................2.4mg
NH 4 NO 3 .....................................................................................1.6mg
Na 2 S·9H 2 O solution...................................................................0.5mL
pH 6.5 ± 0.2 at 25°C
Na 2 S·9H 2 O Solution:
Composition per 10.0mL:
Na 2 S·9H 2 O....................................................................................1.0g
Preparation of Na 2 S·9H 2 O Solution:  Add Na 2 S·9H 2 O to dis-
tilled/deionized water and bring volume to 10.0mL. Mix thoroughly.
Sparge with 100% N 2 . Autoclave for 15 min at 15 psi pressure–121°C.
Neutralize to pH 7.0 with sterile HCl.
Preparation of Medium: Add components, except sulfur and
Na 2 S·9H 2 O solution, to distilled/deionized water and bring volume to
1.0L. Mix thoroughly. Filter through normal filter paper. An iron sedi-
ment will collect in the filter. Gently heat while stirring and bring to
boiling. Boil for 5 min. Cool under an anaerobic gas mixture of 80%
N 2  + 20% CO 2 . Adjust pH to 6.5. Distribute the medium into Hungate
tubes or serum bottles containing finely divided sulfur (0.5% w/v). Seal
the tubes or bottles under the same anaerobic gas used when cooling
the medium. Sterilize the medium at 100°C for 3 hr on 3 consecutive
days. Reduce the medium by adding 10% neutralized Na 2 S·9H 2 O so-
lution to a final concentration of 0.05%. The medium should not give
a heavy black precipitate; if it does the iron sediment was not adequate-
ly removed by filtering in the initial stages and the medium should be
made again, making sure that the iron is removed by filtering.
Use: For the cultivation and maintenance of Thermococcus stetteri
DSM 5262.