Medium H is a growth medium for the culture of algae.

Preparation of 1x Medium H

All solutions are prepared using acid-rinsed glassware and the usual precautions to minimize contamination of unwanted ions.  This medium must be diluted 2X, and Ca²⁺ and Na⁺ must be added to make it a complete medium.  A nitrogen source must also be added except when cells are cultured under dinitrogen fixing conditions.

Stock solutions are stored separately, then are combined as indicated in the table below.

Component                                                                                                    Stock Concentration                      mL stock/500ml                                                                                                                                                                                                                                   

Magnesium Sulfate (MgSO₄-7H₂O, Mol Wt=246.47g )                                   12.5 g/250 mL=0.2 M                                 5

Potassium phosphate dibasic anhydrous (K2HPO4 , Mol Wt=174.18g)           2.6 g/250 mL=0.06 M                                5

HEPES                                                                                                            23.8 g/100 mL=1.0 M                                2

Fe-EDTA                                                                                                              (see below)                                            1

A₅ mnutrients                                                                                                        (see below)                                            1

Sodium Chloride (NaCl, Mol Wt=58.44g)                                                       14.5 g/250 mL=1.0 M                                  1

Calcium Chloride Dihydrate (CaCl₂-2H₂O , Mol Wt=147.01g)                        0.74 g/250 mL=0.02 M                                5

Potassium Nitrate (KNO₃, Mol Wt=101.10)                                                      30.0 g/250 mL=1.2 M                                10

The indicated volume of each stock solution is combined in a 500-ml beaker and diluted into a volume of approximately 450ml.  The pH is adjusted to 7.8 at room temperature, using ~6N KOH.  The final volume is then adjusted to 500ml.  This "2X Minimum Medium H" is stored at 4^oC prior to use.

 Prepare Fe-EDTA Stock Solution as follows:

 To ~150 mL of dH2O add:     H4EDTA (acid form)         2.0 g     (34)   

                                            FeSO₄-7H₂O                  0.8 g     (14)

Titrate the pH to ~7.6 with ~6N KOH.  Then bring the total volume to 200 mL.  If no studies in medium devoid of Na+ are intended, then Na₂H₂EDTA may be substituted for H₄EDTA and the pH may be titrated with NaOH rather than KOH. 

Numbers at right in parentheses for each listed component represents the final micromolar concentration of the component after dilution into Medium H.

 Prepare A₅ + Co Micronutrient Solution as follows:

First prepare a Pre-Stock Solution containing the following:

CuSO₄-5H₂O                    0.32g

Co(NO₃)₂-6H₂O               0.20g

H₂O to bring the total volume to 200 mL.

For the Final Stock Solution add the following components, in the order listed, to a stirred beaker containing ~150 mL of dH₂O.

H₃BO₃                              0.57 g

MnCl₂-4H₂O                    0.36 g

ZnSO4-7H2O                    0.044 g

Na₂MoO₄-2H₂O               0.078 g

Prestock solution            10 mL

H₂O to bring the final volume to 200 mL.

Date Modified: 12/16/2010

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