CulGenex™

LB Agar, Miller Powder

Cat. no. C6000 CulGenex™ LB Agar, Miller, 2L 80gm
Cat. no. C6001 CulGenex™ LB Agar, Miller 500gm
Cat. no. C6002 CulGenex™ LB Agar, Miller 2kg
Cat. no. C6003 CulGenex™ LB Agar, Miller 10kg
Cat. no. C6009 CulGenex™ LB Agar, Miller, 0.5L 6 pouches/pack

INTENDED USE

Hardy Diagnostics CulGenex™ LB Agar, Miller is used for the maintenance and propagation of Escherichia coli used in molecular biology procedures.

This dehydrated culture medium is a raw material intended to be used in the making of prepared media products, which will require further processing, additional ingredients, or supplements.

SUMMARY

LB, or "lysogeny broth", media formulations have been widely used for the cultivation of Escherichia coli since the 1950s, and have become an industry standard in molecular microbiology applications for the preparation of plasmid DNA and the growth of recombinant strains. (3,5-8) LB medium was originally formulated by Giuseppe Bertani and published in 1951 and has since been modified by Miller, Lennox and Luria: the formulations differ in the concentration of sodium chloride, which provides for greater selectivity. (3) LB Agar, Miller medium contains 10gm of sodium chloride; LB Agar, Lennox contains 5gm of sodium chloride; and Luria Agar, Miller contains 0.5gm of sodium chloride. (3,5-8) Low salt formulations, such as those by Lennox and Luria, are ideal for salt-sensitive applications.

Adapted by J.H. Miller, LB Agar is a nutritionally rich medium designed for the growth and culture of pure recombinant strains used in genomic testing. (8) Hardy Diagnostics CulGenex™ LB Agar media formulations are based on the original recipe by Miller and contain casein peptone and yeast extract for amino acids, vitamins and essential minerals. Sodium chloride provides sodium ions for transport and helps maintain osmotic balance. Agar is added as the solidifying agent.

Additional selective agents, such as ampicillin, carbenicillin, chloramphenicol, streptomycin, tetracycline or kanamycin, can be added and are ideal for selective applications; sucrose or glucose can also be added to provide an additional level of selection. The chromogen X-GAL (5-bromo-4-chloro-3-indolyl-galactopyranoside) and supplement IPTG (isopropyl beta-D-thiogalactopyranoside) can be utilized to distinguish lacZ transformed cells, making it easy to differentiate between lac+ (blue) and lac- (white) colonies, if desired.

FORMULA

Gram weight per liter: 40.0gm/L
Yeast Extract 5.0gm
Tryptone 10.0gm
Sodium Chloride 10.0gm
Agar 15.0gm

Final pH 7.0 +/- 0.2 at 25ºC.

* Adjusted and/or supplemented as required to meet performance criteria.

STORAGE AND SHELF LIFE

Store the sealed container(s) containing dehydrated culture medium at 2-30ºC. Dehydrated culture medium is very hygroscopic. Keep container tightly sealed. Protect dehydrated culture media from moisture and light. The dehydrated culture media should be discarded if it is not free-flowing or if the color has changed from its original light tan.

Store the prepared media at 2-8ºC.


PRECAUTIONS


METHOD OF PREPARATION FOR DEHYDRATED CULTURE MEDIA

1. Suspend 40.0gm of the dehydrated culture media in 1 liter of distilled or deionized water (20gm per 500ml).

2. Heat to boiling to dissolve completely.

3. Autoclave at 121ºC. for 15 minutes.

4. Cool to 45-50ºC. Add supplements as needed. Dispense into sterile petri dishes.

LIMITATIONS


MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as autoclave, incinerators, and incubators, etc., are not provided.

QUALITY CONTROL

The following organism is routinely used for testing at Hardy Diagnostics:

Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Escherichia coli
ATCC ® 25922
A 24-48hr 35°C Aerobic Growth

User Quality Control


PHYSICAL APPEARANCE

CulGenex™ LB Agar, Miller powder should appear homogeneous, free-flowing, and very light to light tan in color. The prepared media should appear slightly opalescent, and very light amber in color.

REFERENCES

1. Anderson, N.L., et al. Cumitech 3B; Quality Systems in the Clinical Microbiology Laboratory, Coordinating ed., A.S. Weissfeld. American Society for Microbiology, Washington, D.C.

2. Ausubel, F.M., R. Brent, R.E. Kingston, D.D. Moore, J.G. Seidman, J.A. Smith, and K. Struhl. 1994. Current Protocols in Molecular Biology . Vol. 1. Current Protocols, New York, N.Y.

3. Bertani, G. 1951. Studies on Lysogenesis: The Mode of Phage Liberation by Lysogenic Escherichia coli . J. Bacteriol. ; 62:293-300.

4. Tille, P., et al. Bailey and Scott's Diagnostic Microbiology, C.V. Mosby Company, St. Louis, MO.

5. Lennox, E.S. 1955. Transduction of Linked Genetic Characteristics of the Host by Bacteriophage P1. Virology ; 1:190.

6. Luria, S.E., and J.W. Burrous. 1957. Hybridization Between Escherichia coli and Shigella . J. Bacteriol. ; 74:461-476.

7. Luria, S.E., J.N. Adams, and R.C. Ting. 1960. Transduction of Lactose-Utilizing Ability Among Strain of E. coli and S. dysenteriae and the Properties of the Transducing Phage Particles. Virology ; 12:348-390.

8. Miller, J.H. 1972. Experiments in Molecular Genetics . Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y.

9. Sambrook, J., E.F. Fritsch and T. Maniatis. 1989. Molecular Cloning: A Laboratory Manual , 2nd ed. Cold Spring Harbor Laboratory. Cold Spring Harbor, N.Y.

10. Sambrook and Russell. 2001. Molecular Cloning: A Laboratory Manual, 3rd ed. Cold Spring Harbor Laboratory. Cold Spring Harbor, N.Y.


ATCC is a registered trademark of the American Type Culture Collection.

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