CulGenex™

LB Agar, Lennox Media

Cat. no. G77 LB Agar, Lennox, 15x100mm Plate, 26ml 10 plates/bag

INTENDED USE

Hardy Diagnostics CulGenex™ LB Agar, Lennox Media are recommended for the cultivation of recombinant Escherichia coli strains used in phage production and in molecular studies.

This product is not intended to be used for the diagnosis of human disease.

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 recombinant strains of E. coli . (3,5-8) In general, typical wild type strains of E. coli are capable of growth on minimal media. However, researchers have exploited auxotrophic mutations in strains used for molecular applications, making them dependent upon specific growth factors; one strain in particular, E. coli K-12, is deficient in vitamin B production and is incapable of growth on nutritionally deficient media.

The original recipe for LB medium was formulated by Giuseppe Bertani and published in 1951. (3) LB media have since been adapted by Miller, Lennox and Luria to contain differing concentrations of sodium chloride in order to provide the appropriate osmotic conditions for the strain of interest. LB Agar, Lennox contains half the sodium chloride concentration of LB Agar, Miller and ten times that found in Luria Agar, Miller. (3,5-8) Low salt formulations, such as those adapted by Lennox and Luria, are ideal for salt-sensitive applications.

LB Agar, Lennox was adapted by E.S. Lennox in the mid 1950s and is a nutritionally rich medium designed to contain certain trace elements for the growth and maintenance of pure recombinant strains. Hardy Diagnostics LB Agar, Lennox Media are based on this formulation and contain tryptone and yeast extract for amino acids, vitamins and essential minerals. The moderate amount of sodium chloride (0.5%) provides sodium ions for transport and helps maintain osmotic balance. Agar is the solidifying agent.

There are many factors critical to the transformation process and, in general, E. coli is not naturally transformable. Competency, or the ability to take up extrachromosomal DNA, may be induced by chemical means using divalent cations like calcium or magnesium; divalent cations increase transformation efficiency by increasing cell membrane permeability. Additional agents such as ampicillin, carbenicillin or kanamycin are ideal for selective applications.

FORMULA

Ingredients per liter of deionized water:*

Tryptone 10.0gm
Sodium Chloride 5.0gm
Yeast Extract 5.0gm
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

Storage: Upon receipt store at 2-8ºC. away from direct light. Media should not be used if there are any signs of deterioration (shrinking, cracking, or discoloration), contamination, or if the expiration date has passed. Product is light and temperature sensitive; protect from light, excessive heat, moisture, and freezing.


PRECAUTIONS


PROCEDURE

Consult listed references for recommended test procedures. (2,5,8-10)

INTERPRETATION OF RESULTS

Growth is evident by the formation of isolated colonies or a confluent lawn of bacteria.

LIMITATIONS


MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as loops, swabs, applicator sticks, other culture media, incinerators, and incubators, etc., as well as serological and biochemical reagents, are not provided.

QUALITY CONTROL

The following organisms are routinely used for testing at Hardy Diagnostics:

Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Escherichia coli
ATCC ® 8739
J 1-5 days 35°C Aerobic Growth
Staphylococcus aureus
ATCC ® 6538
J 1-5 days 35°C Aerobic Growth


USER QUALITY CONTROL


Physical Appearance

LB Agar, Lennox should appear slightly opalescent, and 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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