Part A:
Phosphate Buffer (see below)...........200.0 ml
NH4Cl ....................................0.3 g
KNO3 .....................................2.0 g
Sodium benzoate..........................1.0 g
Distilled water........................500.0 ml
Part B:
MgSO4 . 7H2O .............................0.2 g
CaCl2 ...................................10.0 mg
Trace Metals Solution (see below)........1.0 ml
Distilled water........................299.0 ml
Dissolve and autoclave Parts A and B separately.
Combine when cool. Adjust final pH to 8.2. Completely fill screw-capped tubes.
Phosphate Buffer:
K2HPO4 ...................................5.12 g
KH2PO4 ...................................1.5 g
Dissolve in 200 ml distilled water. Adjust pH to 9.0 with KOH.
Trace Metals Solution:
Ferric EDTA (see below).................10.0 ml
ZnSO4 . 7H2O ............................50.0 mg
MnSO4 . H2O .............................50.0 mg
CuSO4 ...................................10.0 mg
Cobalt nitrate..........................10.0 mg
Sodium borate...........................10.0 mg
Sodium molybdate.......................200.0 ml
Distilled water........................100.0 ml
Ferric EDTA:
EDTA....................................17.9 g
KOH......................................3.23 g
FeSO4 . 7H2O ............................13.7 g
Distilled water........................550.0 ml
Dissolve EDTA and KOH in 186 ml distilled water.
Dissolve FeSO4 . 7H2O in 364 ml distilled water.
Mix the two solutions and bubble with air overnight to oxidize the Fe2+ to Fe3+. Store in a dark place.