DM25 Liquid Medium
The basic medium used for propagating the long-term lines is Davis minimal broth (Carlton and Brown 1981*) supplemented with glucose at a concentration of 25 mg per L, which we refer to as DM25. This medium supports a stationary-phase density of about 5 x 107 cells per ml for the founding strain of E. coli B.
Culture volume is 10 ml, and cultures are maintained in 50-ml Erlenmeyer flasks loosely capped with small inverted beakers. Flasks are incubated at 37C and 120 rpm. Cultures are propagated daily by transferring 0.1 ml of each culture into 9.9 ml of fresh medium.
We normally prepare DM25 in 500-ml bottles; and the following recipe makes one 500-ml batch.
|Potassium phosphate (dibasic trihydrate)||3.5 g|
|Potassium phosphate (monobasic anhydrous)||1.0 g|
|Ammonium sulfate||0.5 g|
|Sodium citrate||0.25 g|
Mix thoroughly and autoclave. Immediately after autoclaving, add the following from sterile stock solutions (kept in 100-ml bottles covered by foil):
|Glucose (10%)||125 ul|
|Magnesium sulfate (10%)||500 ul|
|Thiamine (= vitamin B1: 0.2%)||500 ul|
One can, of course, add different concentrations of glucose or other carbon sources as so desired. And one can also make DM agar for plating colonies on glucose or other carbon sources.
Note also that E. coli cannot use citrate to support growth; it serves only as a chelating agent in this medium.
* Carlton, B. C., and B. J. Brown. 1981. Gene mutation. Pp. 222-242 in P. Gerhardt (ed.), Manual of methods for general bacteriology. American Society for Microbiology, Washington, D.C.