HEPES Medium



HEPES Medium Recipe

Modification of the original recipe that changed the buffer of Volvox Medium to accommodate xenic cultures of LB 1662 Glaucosphaera vacuolata. Alkaline medium suitable for axenic and xenic cultures, especially LB 2411 Euglena sp. and LB 1333 Pithophora sp.

 

Directions

For 1 L Total

  1. To approximately 950 mL of dH2O,add each of the components in the order specified (except vitamins) while stirring continuously.
  2. Adjust the pH to 8.2.
  3. Bring total volume to 1 Liter with dH2O.
    • *For 1.5% agar medium, add 15 grams of agar into the flask; do not mix.
  4. Cover and autoclave medium.
  5. When cooled add vitamins.
    • *For agar medium add vitamins, mix, and dispense before agar solidifies.
  6. Store at refrigerator temperature.

# Component Amount Stock Solution Concentration Final Concentration
1 Ca(NO3)2·4H2O (Sigma C 5676) 1 mL/L 11.8 g/100 mL dH2O 0.5 mM
2 MgSO4·7H2O (Sigma 230391) 1 mL/L 4 g/100 mL dH2O 0.16 mM
3 Na2glycerophosphate.5H2O (Sigma G 6501 ) 1 mL/L 5 g/100 mL dH2O 0.16 mM
4 KCl (Fisher P 217) 1 mL/L 5 g/100 mL dH2O 0.67 mM
5 HEPES buffer (Sigma H-3375) 0.94 g/L   3.9 mM
6 P-IV Metal Solution 6 mL/L    
7 Vitamin B12 1 mL/L    
8 Biotin Vitamin Solution 1 mL/L