120 LABORATORY MANUAL OF MICROBIOLOGY 



Exercise 32 



Influence of Nitrate on the Fixation of Nitrogen 



Prepare eight large silica plates containing 2 grams of manni- 

 tol and medium for fixation of nitrogen. Arrange as follows: 



(a) 1 and 2 no nitrate. 



(6) 3 and 4 1.0 mgm. of nitrate nitrogen. 



(c) 5 and 6 5.0 mgm. of nitrate nitrogen. 



(d) 7 and 8 25 . mgm. of nitrate nitrogen. 



Inoculate each with 1 gram of good garden soil. 



Incubate for 7 days at 30°C. 



Dry gel and determine total nitrogen. 



Tabulate results. 



Exercise 33 

 Effect of a Soluble Carbohydrate on Nitrogen Assimilation 



Weigh out two 400- to 500-gram portions of field or garden 

 soil in deep soup plates or shallow earthenware jars. The soil 

 should be thoroughly mixed and sieved. 



(a) Control — untreated. 



(6) Add 2 per cent of mannitol, glucose, sucrose, lactose, or 

 other carbohydrates. 



Mix the carbohydrate thoroughly with the soil by means of a 

 spatula and then add tap water until the moisture content of the 

 soil is about two-thirds saturation. Do not attempt to mix soil 

 and carbohydrate immediately after the water is added. At 

 intervals of about 2 days add more water, sufficient to replace 

 the loss by evaporation and incubate at 28°C. for from 14 to 21 

 days. At the end of this time the carbohydrate should be 

 entirely destroyed. 



Prepare the soil for analysis. When dry, pass it through a 20- 

 mesh sieve, mix thoroughly, and draw a small sample for analysis; 

 about 100 to 150 grams is enough. This smaller sample should 

 be pounded in a mortar until the entire mass passes through a 

 100-mesh sieve. Weigh out from three to six portions of 10 

 grams from each jar into 800-cubic centimeter Kjeldahl flasks 

 and analyze according to the Kjeldahl method not to include 

 nitrates (p. 65). 



