The following is the recipe to prepare ~ 1.0L of minimal salts medium for growth of prototrophic strains of\u00a0E. coli.<\/p>\n
The following are added to 900 mL of autoclaved milliQ water contained in a 2.7L Fernbach flask.<\/p>\n
NOTE: Follow the order of addition given below. Also, be sure to mix the solution thoroughly after each addition.<\/p>\n
1. 100 mL of 10X M9 salts (60g\/L Na2<\/sub>HPO4<\/sub>, 30 g\/L KH2<\/sub>PO4<\/sub>, 5 g\/L NaCl, pH 7.4, autoclave for sterilization).<\/p>\n 2. 10 mL of 100 g\/L NH4<\/sub>Cl, pH 7.4 (use 0.22\u00a0m<\/span>M filter for sterilization (since the ammonia is somewhat volatile, autoclaving is not recommended)).<\/p>\n 3. 2.0 mL of 1.0 M MgSO4<\/sub>\u00a0(autoclave for sterilization).<\/p>\n 4. 12.5 mL 20 % (w\/v) D-glucose (autoclave for sterilization).<\/p>\n 5. 0.2 mL 0.50 M CaCl2<\/sub>\u00a0(autoclave for sterilization).<\/p>\n 6. Appropriate antibiotics, e. g. AMP at 100\u00a0m<\/span>g\/mL (use 0.22\u00a0m<\/span>M filter for sterilization)<\/p>\n 7. 1 mg biotin (added directly)<\/p>\n 8. 0.5 mL 2 mg\/mL thiamine hydrochloride (use 0.22\u00a0m<\/span>M filter for sterilization)<\/p>\n 9. 1.0 mL 15 mg\/mL FeCl2 in 1.0 M HCl (use 0.22\u00a0m<\/span>M filter for sterilization)<\/p>\n