PA: Lac Papillation Agar

Make 10× Minimal A Salts.

10× Minimal A Salts
1L Component MW
80 g Potassium Phosphate (dibasic) K2HPO4 MW 174.18
45 g Potassium Phosphate (monobasic) KH2PO4 MW 136.07
10 g Ammonium Sulfate (NH4)2 SO4 MW 132.08
5 g Sodium Citrate (trisodium, dihydrate) Na3C6H5O7 (H2O)2 MW 294.10
to 1L dH2O  
Autoclave. Salt solution can be stored at room temperature.

Agar Solution
1L Component
15 g Agar
to 490 ml dH2O
Autoclave.

Sugar Solution
1L Component
2 g Glucose
to 400 ml dH2O
Autoclave.

After autoclaving, combine agar and sugar and add:

1L Component
100 ml 10× Minimal A Salts
6 ml 10% w/v (0.083M Mg2+) Magnesium Sulfate, anhydrous MgSO4 (autoclaved) MW 120.37
2.5 ml 0.2% w/v Thiamin (vitamin B1) (filter sterilized)
1 ml 500 mg/ml P-Gal (phenyl β-D-galactoside) (filter sterilized)
1 ml 40 mg/ml X-Gal (5-bromo-4-chloro-3-indolyl β-D-galactoside) (filter sterilized)

Note: The amounts added of some standard components (including agar) are slightly different for this recipe. Volumes have also been altered from the original Miller recipe to use the same stock solutions we have available for making DM plates. The final concentrations are 0.2% glucose, 0.05% P-Gal, and 0.004% X-Gal (w/v).

Sources:

  1. Nghiem, Y., Cabrera, M., Cupples, C.G., Miller, J.H. (1988) The mutY gene : A mutator locus in Escherichia coli that generates GC→TA transversions. Proc. Natl. Acad. Sci. U.S.A. 85:2709-2713.
  2. Miller J.H. (1972) Experiments in Molecular Genetics. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.

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Topic revision: r2 - 19 Feb 2013 - 23:01:24 - Main.AurkoDasgupta
 
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