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HEPES Medium  

Directions 
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.

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 the total volume to 1 L with dH2O.
*For 1.5% agar medium add 15 g 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.

  
#ComponentAmountStock Solution
Concentration
Final Concentration
1Ca(NO3)2·4H2O (Sigma C 5676)1 mL/L11.8 g/100 mL dH200.5 mM
2MgSO4·7H2O (Sigma 230391)1 mL/L4 g/100 mL dH200.16 mM
3Na2glycerophosphate.5H2O (Sigma G 6501 )1 mL/L5 g/100 mL dH200.16 mM
4KCl (Fisher P 217)1 mL/L5 g/100 mL dH200.67 mM
5HEPES buffer (Sigma H-3375)0.94 g/L 3.9 mM
6P-IV Metal Solution6 mL/L  
7Vitamin B121 mL/L  
8Biotin Vitamin Solution1 mL/L  

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