LB

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Contents

Summary

Luria-Bertani Medium (aka L-Broth or LB Medium). (Bertani says LB really stands for lysogeny broth.) LB is a standard growth medium for a variety of bacteria and conditions.

LB contains a milk digest (tryptone) and hence can induce cultures with lac based promoters when they near saturation.

  • "LB ... contains enzymatic digests of the milk protein casein (e.g., tryptone or N-Z-amine) and yeast extract. Since milk is rich in lactose, an inducer of the T7 expression system, variable amounts of residual lactose may be present in different lots of these enzymatic digests. These small amounts of lactose do not promote appreciable induction during log-phase growth, but even minute amounts are sufficient to cause induction on approach to saturation, particularly at lower rates of aeration, which allow induction at lower lactose concentration and promote higher levels of induction." -- Sturdier 2014

Ingredients

  1. 10 g Bacto-tryptone
  2. 5 g yeast extract
  3. 10 g NaCl

For plates also add

  1. 15g agar

See also: Silver: LB Liquid

Note: There are two formulations of LB, Miller and Lennox, that differ in the amount of NaCl. Lennox has less salt, only 5 g/L. The Qiagen miniprep kit recommends LB with 10 g NaCl for highest plasmid yields.

Protocol

  1. Mix dry ingredients and add distilled water up to 1 Liter
  2. Pour into 2 L flask (or greater)
  3. Autoclave (liquid cycle)
    • 250°F, 22psi, 30 minutes

Notes: We do not pH medium when we make it on the fly. However, if it is really important, pH the medium to 7.0 with 5M NaOH (~200µL). Check with pH paper

For plates

  1. Mix dry ingredients and add distilled water up to less than 1 Liter
  2. After dissolved, add agar
  3. Pour into 2 L flask (or greater)
  4. Autoclave (liquid cycle)
    • 250°F, 22psi, 30 minutes
  5. Cool media down to 60-55C (uncomfortable but not painful)
  6. Add appropriate antibiotic
  7. Pour the solution on sterile (???) plates
  8. Let solidify about 1 h, wrap, label (name, date, additive)
  9. keep at 4°C

Source

Adapted From:

J. Sambrook, D.W. Russell, Molecular Cloning: A Laboratory Manual (Cold Spring Harbor Laboratory Press, New York, ed. 3, 2001) pg. A2.2

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