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Methanococcus maripaludis Jones et al.

BAA-1332

Product category
Bacteria
Product type
Extremophile
Strain designation
C6
Type strain
No
Genome sequenced strain
Yes
Isolation source
Salt marsh
Product format
Freeze-dried

Documentation

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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.

Detailed product information

General

Preceptrol
No

Characteristics

Extremophile type
Methanogen
Comments
Genome sequenced strain

Handling information

Medium
Temperature
37°C
Atmosphere
80% H2, 20% CO2
Handling procedure
1. Sterilize the top of the Balch tube by spraying it with 70% ethanol and then flaming the top.

2. Exchange the gas in the test tube for 80% H2 20% CO2, do not pressurize over 5psi.  If the tubes are over pressurized (20 psi.), inoculating the tubes will prove difficult. 

3.  Prepare tubes for inoculation: If the medium is pink (see discussion about resazurin B)  add 2.0 ml of reducing agent (5% Co-enzyme M stock solution) per 100 ml of medium.  Let the medium sit at room temperature for at least 1 hour or until the resazurin becomes colorless, before inoculating.

4. Thaw the frozen vial under a gentle stream of anaerobic gas. Using an anaerobic (see E) 1.0 ml syringe tipped with 22-gauge needle, withdraw the cell suspension from the vial and transfer to the tube of ATCC #1439 medium. Transfer 0.5 ml of the inoculated culture to a second tube of ATCC medium #1439. Pressurize the culture tubes to 20 psi with 80% H2 20% CO2. Plate 0.1 ml of the inoculated culture onto a non-selective medium and incubate the plate aerobically at 37oC.  Incubate culture tubes at 37oC.

5. Growth should be detected in the broth within 1 to 2 days. Growth is enhanced by incubating the cultures with shaking.   No growth should be detected on the aerobic plate.

ANAEROBIC CONDITIONS:

A.         Balch tubes (available from Bellco Glass, Vineland, NJ; are specially designed for anaerobic work and use an aluminum crimp cap to hold a rubber stopper in place. Needles can easily be inserted through the stopper, and the tubes can be pressurized to 2 atm.  Alternatively, serum vials may be used, or screw cap tubes with butyl rubber stoppers, in the latter case the stopper may be removed and the tube placed under a cannula system that dispenses sterile, oxygen free gas for addition of reducing agents or inoculation.

B.         Resazurin is a commonly used redox indicator that is pink when the redox potential is above –50 mv, and colorless when the redox potential is below –110 mv. i.e. highly reducing.  Most strict anaerobes require this low redox potential for optimum growth.

C.         To obtain a fully reduced medium, it is necessary that the medium be anoxic and that a reducing agent be added.  Common reducing agents are sodium sulfide, cysteine, dithiothreitol, titanium citrate and Co-enzyme M (see D).

D.         We suggest adding the reducing agent to the medium at least one hour before the medium is to be inoculated.  Co-enzyme M (mercaptoethanesulfonic acid) (100 X solution):  Dissolve 5.0 g in 100 ml of deionized water.  Distribute into screw cap test tubes, 5–6 ml per tube and seal with rubber stoppers under N2 gas.  Autoclave to sterilize.  Excess tubes can be stored at room temperature for up to 2 months.  Co-enzyme M is a compound produced by many methanogens.  Some methanogens are sensitive to stronger reducing agents such as sodium sulfide.  Co-enzyme M is the standard reducing agent we use when working with methanogens.

E. Syringes can be made anaerobic by one of two methods. 

Handling notes
Cells occur singly and in pairs. The cells exhibited characteristic F420 autofluorescence by epifluorescence microscopy using a low wavelength filter set.

Always use freshly prepared anaerobic medium.  If there is any question about the anaerobic condition of the medium, the it can be reduced with the addition of 5.0%. Co-enzyme M  (2.0 ml per 100 ml of medium).  .

History

Depositors
WB Whitman
Type of isolate
Environmental
Cross references
GenBank CP000867 Methanococcus maripaludis C6, complete genome.

Legal disclaimers

Intended use
This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use.
Warranty

The product is provided 'AS IS' and the viability of ATCC® products is warranted for 30 days from the date of shipment, provided that the customer has stored and handled the product according to the information included on the product information sheet, website, and Certificate of Analysis. For living cultures, ATCC lists the media formulation and reagents that have been found to be effective for the product. While other unspecified media and reagents may also produce satisfactory results, a change in the ATCC and/or depositor-recommended protocols may affect the recovery, growth, and/or function of the product. If an alternative medium formulation or reagent is used, the ATCC warranty for viability is no longer valid.  Except as expressly set forth herein, no other warranties of any kind are provided, express or implied, including, but not limited to, any implied warranties of merchantability, fitness for a particular purpose, manufacture according to cGMP standards, typicality, safety, accuracy, and/or noninfringement.

Disclaimers

This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use. Any proposed commercial use is prohibited without a license from ATCC.

While ATCC uses reasonable efforts to include accurate and up-to-date information on this product sheet, ATCC makes no warranties or representations as to its accuracy. Citations from scientific literature and patents are provided for informational purposes only. ATCC does not warrant that such information has been confirmed to be accurate or complete and the customer bears the sole responsibility of confirming the accuracy and completeness of any such information.

This product is sent on the condition that the customer is responsible for and assumes all risk and responsibility in connection with the receipt, handling, storage, disposal, and use of the ATCC product including without limitation taking all appropriate safety and handling precautions to minimize health or environmental risk. As a condition of receiving the material, the customer agrees that any activity undertaken with the ATCC product and any progeny or modifications will be conducted in compliance with all applicable laws, regulations, and guidelines. This product is provided 'AS IS' with no representations or warranties whatsoever except as expressly set forth herein and in no event shall ATCC, its parents, subsidiaries, directors, officers, agents, employees, assigns, successors, and affiliates be liable for indirect, special, incidental, or consequential damages of any kind in connection with or arising out of the customer's use of the product. While reasonable effort is made to ensure authenticity and reliability of materials on deposit, ATCC is not liable for damages arising from the misidentification or misrepresentation of such materials.

Please see the material transfer agreement (MTA) for further details regarding the use of this product. The MTA is available at www.atcc.org.

References

Curated Citations

Keswani J, et al. Phylogeny and Taxonomy of Mesophilic Methanococcus spp. and Comparison or rRNA, DNA Hybridization, and Phenotypic Methods.. Int. J. Syst. Bacteriol. 46: 727-735, 1996.

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