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C. elegans media

NGM Agar

Mix the following:

NaCl 3 g
Peptone 2.5 g
Agar 17 g
H2O 975 ml

 

Autoclave, cool to 55°C, then add the following (mix after each addition):

Cholesterol (5 mg/ml in EtOH) 1 ml
1 M CaCl2 1 ml
1 M MgSO4 1 ml
1 M Potassium Phosphate pH 6.0 25 ml

 

RNAi plates

To regular NGM medium add (per L):

50 mg Ampicillin
240 mg IPTG (1 mM)

 

Egg plates

(Adapted from Methods in Cell Biology 48, page 515)

Egg plates can be used to grow large amount of worms on plates, and are especially usefull to grow unc-119 mutants. The egg yolks support rapid bacterial growth.

  1. In a sterile bottle, mix the following:
    • Yolk of 10 eggs (sterilize eggs by washing them in 1% SDS)
    • LB-medium to 500 ml
  2. Heat the mixture to 60°C for 10 min (to inactivate the lysozyme).
  3. Add 50 ml of an O/N OP50 culture.
  4. Dispense onto 2% NGM-agar plates (~4 ml for 10 cm plates and 8 ml for 15 cm plates).
  5. Dry the plates, and keep at 4°C.

 

M9 Buffer

Mix the following and autoclave:

KH2PO4 3 g
Na2HPO4 6 g
NaCl 5 g
1 M MgSO4 1 ml
H2O 1 L

 

S Medium

Mix the following sterile solutions:

S Basal 1 L
1 M Potassium Citrate pH 6.0 10 ml
Trace Metals Solution 10 ml
1M CaCL 3 ml
1M MgSO4 3 ml

 

S Basal

Mix the following and autoclave:

NaCl 0.1 M
Potassium Phosphate pH 6.0 0.05 M
Cholesterol (5 mg/ml in EtOH) 1 ml
H2O 1L

 

Trace Metals Solution

Mix the following and autoclave. Keep in the dark.

Disodium EDTA 1.86 g 5 mM
FeSO5·7H2O 0.69 g 2.5 mM
MnCl2·4H2O 0.20 g 1 mM
ZnSO4·7H2O 0.29 g 1 mM
CuSO4·5H2O 0.025 g 0.1 mM
H2O 1L

 

Freezing Solution

Mix the following and autoclave:

NaCl 5.85 g 10 mM
KH2PO4 6.8 g 20 mM
Glycerol 300 g 30% w/v
1M NaOH 5.6 ml 5.6 mM
H2O to 1L

After autoclaving, add:

0.1 M MgSO4 3 ml

 

1M Potassium Phosphate pH 6.0

  • Make 1M solutions of K2HPO4 and KH2PO4
  • Mix the two in the following amounts:
    • 132 ml K2HPO4
    • 868 ml KH2PO4

In theory this should yield a pH of 6.0. In practice however, the final pH can deviate from this. For a more accurate final pH, start with 868 ml of KH2PO4, then add K2HPO4while measuring the pH until it reaches 6.0.

 

Egg-Salts buffer

Mix the following and autoclave:

Hepes pH 7.4 0.025 mM
NaCl 118 mM
KCl 48 mM
MgCl2 2 mM
CaCl2 2 mM