Ruminococcus bromii Moore et al.
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ATCC determines the biosafety level of a material based on our risk assessment as guided by the current edition of Biosafety in Microbiological and Biomedical Laboratories (BMBL), U.S. Department of Health and Human Services. It is your responsibility to understand the hazards associated with the material per your organization’s policies and procedures as well as any other applicable regulations as enforced by your local or national agencies.
ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.
2. Perform all steps under anaerobic conditions (see below).
3. Aseptically transfer 0.5 ml of #1016 broth to the vial and rehydrate the pellet. Transfer the suspension back into the broth tube. Inoculate a plate of a non-selective medium such as Tryptic Soy, Nutrient, or blood agar with 0.1 ml of the cell suspension.
4. Seal the tube with a rubber stopper and incubate anaerobically at 37oC. Incubate the plate(s) aerobically as a purity check.
5. After one or two days, growth should be evident as indicated by turbidity that is easily resuspended through out the broth by inverting the tube. Once growth has been established the culture should be transferred to fresh broth every 48 - 72 hours.
6. This culture is sensitive to oxygen when initially rehydrated; therefore steps should be taken to avoid exposure to oxygen. When the culture exhibits good growth it will remain viable for up to 1 week if stored at 4oC under anaerobic conditions.
· Tubes of media are placed under a gassing cannula system hooked to a source of oxygen free gas.
· All transfers are performed while the test tubes are on the cannula system with a gentle stream of oxygen free gas (97% CO2 3%H2) flowing through the system.
· As the test tubes are removed from the cannula system each is sealed with butyl rubber stopper thus maintaining the anaerobic headspace.
Always use freshly prepared anaerobic medium. If there is any question about the anaerobic condition of the medium, the medium can be reduced with the addition of 1.5% cysteine
(2.0 ml per 100 ml of medium).
Other commonly used reducing agents are sodium sulfide, cysteine, dithiothreitol, and titanium citrate
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