M9

sterile H₂O (cooled to 50°C or less), to 750 ml

5x M9 salts, 200 ml

1 M MgSO₄, 2 ml

20% solution of the appropriate carbon source (e.g., 20% glucose), 20 ml

1 M CaCl₂, 0.1 ml

sterile deionized H₂O, to 980 ml

For solid medium, please see Media Containing Agar or Agarose

If necessary, supplement the M9 minimal medium with stock solutions of the appropriate amino acids and vitamins.

Prepare the MgSO₄ and CaCl₂ solutions separately, sterilize by autoclaving, and add the solutions after diluting the 5x M9 salts to 980 ml with sterile H₂O. Sterilize the glucose by passing it through a 0.22-μm filter before it is added to the diluted M9 salts.

When using E. coli strains that carry a deletion of the proline biosynthetic operon [Δ(lac-proAB)] in the bacterial chromosome and the complementing proAB genes on the F' plasmid, supplement the M9 minimal medium with the following:

0.4% (w/v) glucose (dextrose)

5 mM MgSO₄•7H₂O

0.01% thiamine