Common Worm Media and Buffers   

Abstract

Materials and Reagents

1. Agar, peptone (BD Biosciences)
   
2. Cholesterol (Sigma-Aldrich)
   
3. Streptomycin (Sigma-Aldrich)
   
4. Nystatin (Life Technologies, Gibco^®)
   
5. Bleach (Clorox)
   
6. Potassium phosphate
   
7. Clorox bleach
   
8. NaCl
   
9. CaCl₂
   
10. MgSO₄
    
11. EtOH
    
12. FeSO₄.7H₂O
    
13. Na₂EDTA
    
14. MnCl₂.4H₂O
    
15. ZnSO₄.7H₂O
    
16. CuSO₄.5H₂O
    
17. KH₂PO₄
    
18. Na₂HPO₄
    

Equipment

1. 60 x 15 mm plate
   
2. Plastic boxes
   

Recipes

1. Nematode growth medium (NGM) agar: For the maintenance of worms.
   For 1 liter medium
   3 g NaCl
   17 g agar
   2.5 g peptone
   1 ml cholesterol (5 mg ml^-1 in 95% EtOH)
   975 ml H₂O
   Autoclave, and then add the following sterile solution (autoclaved)
   1 ml 1 M CaCl₂
   1 ml 1 M MgSO₄
   25 ml 1 M potassium phosphate (pH 6) (to avoid precipitation, mix between addition of MgSO₄ and potassium phosphate)
   To make 1 M potassium phosphate (pH 6): For 1 liter, dissolve 136.1 g KH₂PO₄ in about 800 ml dH₂O, then adjust to pH 6.0 with solid KOH (approx 15 g) before bringing up to volume. Make 100 ml aliquots and autoclave.
   Need to add streptomycin (300 ng ml^-1) if plate is used for seeding bacterial food E coli OP50-1. Typically pour 60 x 15 mm plate and store NGM plates in plastic boxes with covers at room temperature.
   
2. S-basal medium (adapted from the Kim Lab at Stanford) : For liquid culture of worms.
   For 1 liter medium
   5.8 g NaCl
   50 ml 25 ml 1 M potassium phosphate (pH 6)
   1 ml cholesterol (5 mg ml^-1 in 95% EtOH)
   950 ml dH₂O
   Autoclave, and then add the following sterile solution (autoclaved)
   3 ml 1 M CaCl₂
   3 ml 1 M MgSO₄
   10 ml trace metals solution
   10 ml 1 M potassium citrate (pH 6.0)
   10 ml 100x Nystatin (antifungal agent, keep in freezer; do not have to add it all the time).
   To make 500 ml trace metals solution
   0.346 g FeSO₄.7H₂O
   0.930 g Na₂EDTA
   0.098 g MnCl₂.4H₂O
   0.144 g ZnSO₄.7H₂O
   0.012 g CuSO₄.5H₂O
   Sterilize by autoclaving. Keep in dark (wrap in foil).
   To make 100 ml of 1 M potassium citrate: dissolve 21.02 g citric acid, monohydrate in 80 ml and adjust to pH 6.0 with solid KOH (approx 17g) before bringing up to volume.
   
3. Worm M9 buffer
   3 g KH₂PO₄
   6 g Na₂HPO₄
   5 g NaCl
   Add H₂O to 1 liter. Sterilize by autoclaving.
   After solution cools down, add 1 ml autoclaved/sterile 1 M MgSO₄.
   
4. 100 ml 2x worm lysis solution: For worm egg prep
   50 ml ddH₂O
   10 ml 10 M NaOH
   40 ml Clorox bleach
   Make fresh and store at 4 °C up to one week.