156 SOIL BACTERIOLOGY 



Mercuric Chlorid in Vacuum : 



1. Set up the apparatus shown in Fig n, using heavy 

 walled bottles (milk) and heavy steam-proof rubber connec- 

 tions. 



2. Fill flask B with 0.25 per cent, solution of mercuric 

 chlorid and C with distilled water. 



3. After the whole apparatus is connected, the clamps 

 between bottles B-D and C-D fastened, sterilize in the 

 autoclave at 10 pounds' pressure for fifteen minutes. 



4. Cool to 40 C., connect to a vacuum pump, and place 

 the seed in flask D. If the seed are small, place a layer of 

 cheese-cloth over the mouth of bottle before inserting 

 stopper. The seed should be thoroughly clean before 

 sterilizing. It is well to wash with 60 to 70 per cent, 

 alcohol. 



5. By means of the vacuum pump draw the mercuric 

 chlorid from B to D; then close the screw-clamp and 

 exhaust D for three to five minutes. This should remove 

 the air particles from around the seed coats and allow 

 the disinfectant to come in direct contact with the 

 seed. 



6. At the end of this time invert D and withdraw the 

 mercuric chlorid solution. Now run in a small amount of 

 sterile water from C, shake vigorously, empty, and repeat 

 this process three or four times. 



7. Remove some of the seed to sterile Petri dishes and 

 pour over them a layer of bouillon agar. 



8. After the agar hardens, invert and place in the in- 

 cubator at 20 to 25 C. In two or three days the seed 

 should germinate. If bacteria or molds are present, they 

 may be readily noted on the agar. 



Hutchinson and Miller, Jour. Agr. Sci., vol. iii, p. 185, 1908. 



