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From 2010.igem.org

Jeremy's Notebook

Growing Broth for Pseudomonas

Composition of Medium * 100 mg Naphthenic acids /L

(Modified Bushnell-Has medium) * 1000 mg KH2PO4 /L * 1000 mg Na2HPO4 /L * 500 mg NH4NO3 /L * 500 mg (NH4)2SO4 /L * 200 mg MgSO4 ∙ 7H2O /L * 20 mg CaCl2 ∙ 2H2O /L * 2 mg FeCl3 /L * 2 mg MnSO4 ∙ 2H2O /L pH of Bushnell-Has Medium: 7.0 Protocol * Prepare naphthenic acids stock * Mix 1 g naphthenic acid /L in dilute NaOH. * Adjust pH of the solution to 10-11 to dissolve naphthenic acids into the solution as sodium salts * Making the Growing Broth * Mix 20 ml naphthenic acid stock with 170 ml sterilized water in 500 ml erlenmeyer flask. * Prepare 10 ml Bushnell-Has medium in vial * Mix arbitrary amount of pseudomonas strain in B-H medium * Transfer medium to erlenmeyer flask * Incubation (aerobic conditions) * Incubate stock at room temperature (20 °C) on a shaker at 200 rpm.