Lysogeny broth, commonly shortened to LB both for the broth and agar, and often incorrectly called "Luria broth" or "Luria-Bertani medium", is the most commonly used rich medium for bacterial culture, especially for E. coli. It is a solution of tryptone, yeast extract, and NaCl. The main carbon source is the oligopeptides in tryptone, the result of digestion of cow milk casein with the protease trypsin. The amino acids from these peptides are utilized as primary carbon sources in order of preference. Growth in LB does not acidify the medium as with growth with sugar carbon sources; it alkalifies it to pH 9 at saturation(1) at which point exhaustion of a carbon source arrests growth (not pH extreme)(1). Yeast extract is a complex, widely used hydrolysate of yeasts that provides amino acids/peptides, nucleotides, water-soluble metabolic cofactor "B vitamins", some carbohydrates, and other growth enhancers, which can enable growth of diverse microbes with differing nutrient needs. Yeast extract content was found to be the main determining factor for plasmid yield in E. coli, resulting in a higher yeast extract version of LB called LB30.
LB has limitations; no conclusions about E. coli physiology ought to be made in LB(2), but rather in a rationally-designed defined medium, like M9 or MOPS minimal(3).
- Combine the following in an appropriate sized flask or bottle:
For broth:
- 25 g/L LB powder
- or from separate components:
- 10 g/L tryptone
- 5 g/L yeast extract
- 10 g/L NaCl for LB-Miller (common). 5 g/L for LB-Lennox. 0.5 g/L for LB-Luria.
For agar:
- 40 g/L LB agar powder
- or from separate components:
- 10 g/L tryptone
- 5 g/L yeast extract
- 10 g/L NaCl for LB-Miller (common). 5 g/L for LB-Lennox. 0.5 g/L for LB-Luria.
- 15 g/L agar
For LB30: Add an additional 25 g/L yeast extract to any recipe.
- Close and shake to partially combine. Flasks can be closed with parafilm.
For agar dishes (plates), adding a stir bar is advised for quicker and uniform cooling, unless using a 50–55° water bath. - Sterilize by autoclaving for 20 min on the liquid cycle.
- Cool the medium to 50–55°C. Add any antibiotics or other thermosensitive additives only when the vessel's temperature has fallen to 50–55°C. Swirl thoroughly to distribute additives if not using a stir bar.
- Cooling can be sped and made uniform by stirring with a stir bar, further sped by a cold water bath.
- For agar, monitor temperature closely using a tape thermometer held against the vessel surface or an infrared thermometer. Plates ought to be poured soon after medium reaches 50–55°C to avoid congealing in the vessel.
- Polystyrene Petri dishes may deform if agar is much hotter.
- For agar, dispense ≈20 mL into each petri dish. Work quickly to avoid premature agar congealing.
- Pouring is fastest, but less accurate: stop pouring when ≈⅔ dish is covered; then let the remainder fill, swirling the dish or stack of dishes if needed.
- For more accuracy, though slower, use a 50 mL pipette to dispense.
- Stacks of plates can be efficiently filled from the bottom up: fill a dish with one hand while holding its lid below the remainder empty stack above it in the other hand.
- Bubbles can be eliminated by very briefly passing a flame over the surface.
- Sezonov, Guennadi, Danièle Joseleau-Petit, and Richard D'Ari. "Escherichia coli Physiology in Luria-Bertani Broth." Journal of bacteriology 189.23 (2007): 8746-8749.
- Nikaido, Hiroshi. "The Limitations of LB Medium." Small Things Considered, http://schaechter.asmblog.org/schaechter/2009/11/the-limitations-of-lb-medium.html.
- Neidhardt, Frederick C., Philip L. Bloch, and David F. Smith. "Culture Medium for Enterobacteria." Journal of bacteriology 119.3 (1974): 736-747.
- Wood, Whitney N., et al. "Enhancing yields of low and single copy number plasmid DNAs from Escherichia coli cells." Journal of microbiological methods 133 (2017): 46-51.