5x M9 Salts
- Using premixed Difco 5x right now
M9 (for 1 L)
- Add 100 uL 1M CaCl2 to 750 mL autoclaved, warm water (see note)
- Add 2 mL 1M MgSo4
- Add 200 mL 5x M9 salts
- Add carbon source
- Add any other supplements
- Add autoclaved water to 1L
- Make sure autoclaved water for diluting the M9, etc. is warm. M9 salts bottle suggests 40-50 deg C. I keep a bottle of autoclaved water for preparing this media in the 65 deg C incubator, so it's ready when I need it.
- Mix between each component especially the CaCl2 and the MgSO4
- Add CaCl2 to the water first.
M9 Agar, 150 mL
- Add 2.25g of agar to H20 to 100 mL dH2O
- After autoclaving, swirl to mix evenly and "temper" at room temperature until you can place your hand on the flask for 2 second
- Add 0.3 mL of 1M MgSO4, .015mL of 1M CaCl2 + carbon source of choice
- Add 30mL 5X M9 salts solution
- Add H2O to 150 mL
- DO NOT ADD 5x M9 Salts to Agar before autoclaving, otherwise you'll get gross brown agar that nothing will grow on.