What is LB medium?
LB stands for Luria-Bertani, or Lysogeny Broth, and is the most widely used medium for growing E. coli in molecular biology. It has only three ingredients: tryptone for amino acids and peptides, yeast extract for vitamins and growth factors, and sodium chloride for osmotic balance. The three standard versions differ only in the amount of salt.
LB broth formulations (per litre)
| Ingredient | LB Miller | LB Lennox | LB Luria |
|---|---|---|---|
| Tryptone | 10 g | 10 g | 10 g |
| Yeast extract | 5 g | 5 g | 5 g |
| NaCl | 10 g | 5 g | 0.5 g |
| Total (dehydrated medium) | 25 g/L | 20 g/L | 15.5 g/L |
| pH after sterilisation (25 °C) | 7.0 ± 0.2 | 7.0 ± 0.2 | 7.0 ± 0.2 |
- Miller: the general-purpose version with the most salt, used for routine E. coli culture and plasmid preparation.
- Lennox: half the salt, chosen when working with salt-sensitive antibiotics such as blasticidin or zeocin.
- Luria: almost no salt, for special applications.
For LB Agar, add 15 g/L agar to any of the three.
How to prepare LB broth
- Weigh 25 g of dehydrated LB Miller, or 10 g tryptone, 5 g yeast extract and 10 g NaCl.
- Add about 900 mL deionised water and stir until dissolved. LB dissolves without heating.
- Make up to 1 L. No pH adjustment is needed; a correct formulation lands near 7.0 after sterilisation. If you must adjust, use 1 N NaOH to 7.0 to 7.5.
- Dispense into bottles no more than two-thirds full, caps loosened.
- Autoclave at 121 °C for 15 minutes.
- Cool to room temperature before use. Store at 4 °C in the dark and use within two weeks.
How to prepare LB agar
- Add 25 g dehydrated LB plus 15 g agar (or 40 g of a ready LB Agar powder) to 1 L deionised water.
- Heat to boiling with stirring until the agar is completely dissolved, otherwise plates will be uneven.
- Autoclave at 121 °C for 15 minutes.
- Cool to 50 to 55 °C (the bottle can be held by hand) before adding antibiotics; higher temperatures inactivate them.
- Pour 20 to 25 mL per plate, leave lids ajar until set, then store inverted at 4 °C.
When to add antibiotics
Always after sterilisation, once the medium is below 55 °C. Never autoclave antibiotics with the medium. Typical working concentrations: ampicillin 100 µg/mL, kanamycin 50 µg/mL, chloramphenicol 25 µg/mL, tetracycline 10 µg/mL (protect from light).
Frequently asked questions
The medium darkened or formed a precipitate after autoclaving. Usually over-sterilisation, which drives the Maillard reaction between sugars and amino acids, or hard water. Shorten the cycle and use deionised water. See the article on media turning dark or cloudy after autoclaving.
The agar did not set. The agar was not fully dissolved, or the pH was too low. Boil to a clear solution before pouring. See the article on agar not solidifying.
Growth is slow. First confirm whether you are using Miller or Lennox, then look at the tryptone. Tryptones from different sources differ in amino acid profile; compare growth curves when changing supplier. See LB Broth Miller vs Lennox.
Can LB be used for yeast? Yeast will grow, but YPD is the better choice. LB was designed for bacteria and yeast needs more glucose.
Buy dehydrated medium or weigh your own?
Weighing the three ingredients is cheapest, but every weighing adds error and batches drift. Dehydrated culture medium is pre-blended; add water and it is ready, which suits routine use and shared labs. STBIO MEDIA supplies LB Broth Miller, LB Broth Lennox, LB Agar and the tryptone and yeast extract to make your own. Specifications and pack sizes are on each product page.