M9 Minimal Media (500mL)

1. In 500mL glass bottle autoclave approximately 325mL of water with 7.5g agar.

2. To this agar mixture add 100mL 5X M9 Salts and then add sterile water up to 500mL.

3. Add 1mL 1M MgSO_4.

4. Add 50μL 1M CaCl₂.

5. Add 20mL 20% glucose (or other carbon source).

You can also add:

Amino acids to a final concentration of 50 μg/mL

D-Biotin to a final concentration of 1 μg/mL

Notes:

1. Make 1M MgSO₄ and 1M CaCl₂ stocks separately and autoclave.

2. Filter sterilize (DON’T AUTOCLAVE) glucose solution before adding to media.

3. Prepare amino acids at 5mg/mL (100X) and filter sterilize.

4. Prepare biotin stock at 0.1g/L (100X) and filter sterilize.