504 APPENDIX 



The nitrilotriacetic acid is dissolved and neutralized with 

 KOH (about 7.3 g), after which the rest of the ingredients are 

 added. The pH is adjusted to 6.6-6,8 before making to volume, 

 Metals "44" contains, per 100 ml: 



Ethylenediamine tetraacetic acid 250,0 mg 



ZnS04 • 7 H2O 1095,0 mg (250 mg Zn) 



FeS04 • 7 H2O 500,0 mg (100 mg Fe) 



MnS04 • H2O 154.0 mg (50 mg Mn) 



CUSO4 • 5 H2O 39.2 mg (10 mg Cu) 



Co(N03)2 • 6 HgO 24.8 mg (5 mg Co) 



Na2B407 • 10 H2O 17.7 mg (2 mg B) 



A few drops of sulfuric acid are added to retard precipitation. 



In order to prepare 1 liter of the complete medium, 20 ml each of 

 solutions 1, 2 and 3 are mixed and diluted with distilled water, after 

 which 1.0 g of vitamin-free casamino acids (Difco) is added. A copious 

 precipitate forms when this medium is autoclaved, but it redissolves 

 on cooling. The casamino acids may be replaced by a mixture of L- 

 glutamic acid (0.1% final concentration) and sodium acetate • 3 H2O 

 (0.1% final concentration). 



Incubation temperature: 30°C. 



B. Medium S. After Lascelles, Biochem. J., 62, 78 (1956). 

 Sodium- L-glutamate monohydrate 3.8 g 

 DL-malic acid 2,7 g 

 KH2PO4 500.0 mg 

 K2HPO4 500.0 mg 

 (NH4)2HP04 800.0 mg 

 MgS04 • 7 H2O 200.0 mg 

 CaCl2 40.0 mg 

 Nicotinic acid 1.0 mg 

 Thiamine • HCl 1.0 mg 

 Biotin 10.0 //g 

 Distilled water to 1 liter. 



The pH is adjusted to 6.8 with 1 N NaOH before autoclaving for 10 

 minutes at 10 lbs,/sq, in. 



C. Medium MS. After Lascelles, Biochem. J., 72, 508 (1959). 

 Medium S, plus: MnS04, 5 /WM; sodium- L-glutamate, 0.01 M. 



D. Ormerod, Ormerod and Gest (1961) medium for R. nibnon, sup- 

 plemented with 300 mg (Difco) yeast extract per liter, 



E. Keane cl al . medium (1963) for Rps . palustris. 



