Standard Microbiology Reagents

Agarose gel loading buffer

0.25% (w/v) bromophenol blue, 0.25% (w/v) xylene cyanol FF, 30% (v/v) glycerol.

50x Denhardt’s solution

0.5 g each of bovine serum albumin (Sigma cat. no. A-3425), Ficoll (Sigma cat. no. F-2637) and polyvinylpoly pyrrolidine (Sigma cat. no. P-6755) were dissolved in 50 ml distilled water. The mixture was filter sterilised through a 0.2 µ filter, then stored at -20°C until required.

LB medium (Luria-Bertani; liquid broth)

1 g Bactotryptone (Oxoid, cat. no. L42), 0.5 g Yeast extract (Oxoid, cat. no. L21) and 1 g sodium chloride were dissolved in 500 ml distilled water, adjusted to pH 7.0 with 0.5 M sodium hydroxide and sterilised by autoclaving.

LB medium (agar)

500 ml of LB (liquid broth) media was prepared as described above. Before autoclaving, 1.5 g Bacteriological agar (Oxoid cat. no. L11) was added to the solution.

10x MOPS buffer

For 500 ml of 10x MOPS buffer (0.4 M MOPS, 0.1 M sodium acetate and 10 mM EDTA), 41.85 g MOPS, 4.1 g sodium acetate and 0.94 g EDTA were added to 400 ml distilled water and the pH adjusted to 7.2 with sodium hydroxide. 250 µl DEPC were added to the solution (to inactivate RNAses), which was then made up to 500 ml with distilled water. The solution was sterilised by autoclaving.

SOC media

The constituents of the media were dissolved in distilled water to give a final concentration of 20 mM glucose, 2.5 mM potassium chloride, 20 mM magnesium chloride, 20 mM magnesium sulphate, 10 mM sodium chloride, 2% (w/v) tryptone, 0.5% (w/v) yeast extract. The solution was sterilised by autoclaving.