ATCC medium: 1442 Methanoculleus thermophilus medium
MgCl2 . 6H2O .............................4.0 g
CaCl2 . 2H2O .............................0.075 g
NaCl....................................11.7 g
K2HPO4 ...................................0.14 g
KH2PO4 ...................................0.14 g
NaHCO3 ...................................4.0 g
Yeast Extract (BD 212750)................2.0 g
Trypticase Peptone (BD 211921)...........6.0 g
Resazurin................................1.0 mg
L-Cysteine . HCl.........................0.5 g
Na2S .....................................0.25 g
Wolfe's Vitamin Solution (see below)....10.0 ml
Sodium formate...........................5.0 g
Distilled water to.......................1.0
Add all constituents except cysteine and sodium sulfide to the distilled water. Bubble the solution with oxygen-free N2 and CO2 (80-20%) gas for 20 minutes. Adjust the pH to 7.2 with 2 M KOH and add the cysteine and sodium sulfide. Tube the medium under N2 and CO2 (80-20%) in either serum tubes or bottles. After sterilization the medium may precipitate out; allow it to cool and mix to bring the precipitate back into solution.
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