Nematode Growth Medium (NGM)

• 1. For solid NGM, mix 3 g of NaCl, 2.5 g of peptone, and 20 g of agar and bring to 1 L with H₂O. (For liquid NGM, prepare without agar.)

• 2. Autoclave for 1 h.

• 3. Let cool for 1 h in a 55°C water bath.

• 4. Add 1 mL of cholesterol (5 mg/mL in ethanol), 1 mL of 1 m CaCl₂, 1 mL of 1 m MgSO₄, and 25 mL of 1 m (pH 6.0) KPO₄, mixing after each addition.

• 5. Using an automated plate pourer (peristaltic pump), pour the medium into sterile plastic plates to set. During pouring, keep the NGM on a hot plate with stirrer to prevent the medium from solidifying.

• 6. Store NGM plates at 4°C until use. Seed NGM plates with bacteria the day after pouring. Warm the plates to room temperature before adding the worms. (For liquid NGM, store at room temperature for up to 1 mo. Regularly check for excess cloudiness to ensure bacterial contamination is absent; this is particularly important for liquid RNAi cultures.)

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