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Originally from  Tartoff, et al. (1987), printed in Cold Spring Harbor Protocol. Characterized with LB in Danquah & Forde (2007).  

Plasmid yield for TB is ≈10 µg/mL for pUC19, fourfold more than LB, but at 65% the medium cost per amount plasmid, according to Danquah & Forde. Shyam finds that E. coli TG1 (parent of Turbo) saturates at OD600 ≈ 18–20 in and at OD 600 ≈ 6 in LB.

Terrific Broth (TB)

  • 12 g/L tryptone (1.2% m/V)
  • 24 g/L yeast extract (2.4% m/V)
  • 4 mL/L glycerol (4.34 mM, 0.4% V/V)
  • 17 mM KH2PO4, 2.31 g/L
  • 72 mM K2HPO4, 12.54 g/L
  • (opt.) 15 g/L agar (1.5% m/V)
  • (opt., nontrad.) 5–10 mM 1 M MgSO 4  or MgCl 2

It was originally suggested to separately autoclave phosphate buffer and agar. It was recently found that phosphate and agar react to form H2O2 , but  E. coli are less, if at all, significantly impacted due to catalase expression. Commercial mixes do not separate the two, yet instruct autoclaving. There is also reason for separately autoclaving the glycerol, as glycerol partakes in Maillard reactions with tryptone, though probably less so than sugars do due to its lack of carbonyls. Maillard reactions sequester nutrients and form products that decompose into growth-inhibitory compounds.

Magnesium is deficient in LB. It has been found that in peptone and tryptone broths, Mg 2+ is necessary for robust growth, especially when glucose is provided. There is a substantial growth improvement with 1 mM MgCl 2 even in phosphate-buffered tryptone broth, elucidating the insufficiency of contaminant Mg 2+ in phosphate salts, though phosphate does bring in substantial contaminant trace minerals. 1–10 mM MgSO4 or MgCl2 can be supplemented to any peptide broth for best growth and minimal cell stress.

2×YT Medium

  • 16 g/L tryptone (1.6% m/V)
  • 10 g/L yeast extract (1.0% m/V)
  • 5 g/L NaCl (0.5% m/V)
  • (opt.) 15 g/L agar (1.5% m/V)
  • (opt., nontrad.) 5–10 mM 1 M MgSO 4  or MgCl 2

 

Recovery Medium:

Shyam  rationalizes that TB or 2×YT supplemented with 0.4% V/V glucose and MgCl2 and/or MgSO 4  to a final 10–20 mM would be just as good as SOC for transformation recovery. SOB/SOC, has 10 mM MgCl 2  and 10 mM MgSO 4  to help rebuild the cation-associated E. coli outer membrane LPS, and glucose is an ideal carbon source.

Procedure for 1 L TB

  1. Dissolve and autoclave on liquid cycle, 20 min, 15 psi:
    1. 12 g tryptone
    2. 24 g yeast extract
    3. 4 mL glycerol (or 8 mL 50% glycerol)
      (or add after autoclaving) 
    4. (opt.) 15 g agar
    5. (opt.) 5–10 mL 1 M MgSO 4  or MgCl 2
    6. final 900 mL diH2O
  2. Prepare or obtain 100 mL 10× TB buffer (0.17 M KH2PO4 0.72 M K2HPO4)
    Dissolve and autoclave on liquid cycle, 15 min, 15 psi:
    1. 2.31 g KH2PO4
    2. 12.54 g K2HPO4
    3. ≈90 mL diH2O, adjusted to 100 mL final
  3. When 900 mL nutrient solution has cooled ≤60°C, add 100 mL sterile buffer.

 

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