ARTIFICIAL CULTURES FOR INOCULATION OF LEGUMES 73 



3. Incubate at 28 C. for three weeks. 



4. Then analyze for total nitrogen. 



Exercise 39 

 Production of Gum by Bacillus Radicicola 



1. Inoculate two tubes of different media (42 and 47) 

 with Bacillus radicicola, and at the same time leave two 

 tubes uninoculated. 



2. Incubate for four to five weeks at 28 C., and test for 

 gum. 



3. Add 10 c.c. of alcohol (95 per cent.) or 5 c.c. of acetone 

 to each tube. 



4. Note the precipitation of gum. 



Exercise 40 

 Artificial Cultures for the Inoculation of Legumes 



A. i. Inoculate one bottle of mannit agar (m. 42) and 

 one of mannit solution with a pure culture of legume 

 bacteria. Use 5oo-c.c. bottles with flat sides. In the 

 liquid culture take 300 c.c. of the medium; in the agar 

 culture, 100 c.c. Dilute a young legume culture of the 

 desired organism with 5 c.c. of sterile water. Shake 

 thoroughly and pipet i-c.c. portions into the bottles of 

 liquid and solid media. Be sure that the inoculum covers 

 the entire surface of the agar. 



2. Incubate the cultures at 28 C. for four to ten days. 



3. Determine the number of bacteria in each bottle. The 

 liquid culture may be treated as follows: Shake thoroughly, 

 remove i c.c. to a gg-c.c. water blank, and continue the 

 dilution to i : 1,000,000 and i : 10,000,000. Pour mannit 

 agar plates. 



