Methanosarcina mazei (Barker) Mah and Kuhn

1. Open thawed vial according to enclosed instructions or visit www.atcc.org for instructions.
2. Under anaerobic conditions aseptically transfer the entire contents to a 5-6 mL tube of #2467 broth. Additional test tubes can be inoculated by transferring 0.5 mL of the primary broth tube to these secondary broth tubes. Best practice dictates the use of pre-reduced media.
3. Add acetate to a final concentration of 50 mM.
4. Incubate in an anaerobic atmosphere at 37°C for 5-7 days. Incubate one agar plate aerobically at 37°C to check for contamination.

• Anaerobic conditions for transfer may be obtained by the use of an anaerobic gas chamber or placement of test tubes under a gassing cannula system connected to anaerobic gas.

• Loose screw caps on test tubes in an anaerobic chamber
• Loose screw caps on test tubes in an activated anaerobic gas pack jar
• Use of sterile butyl rubber stoppers on test tubes so that an anaerobic gas headspace is retained

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