EFFECT OF CONJUGATION 345 



(non-conjugants) or September 18 (conjugants), a period of 24 

 (or 26) days. 



Culture methods necessary to secure uniformity of conditions. 

 In order that the conditions should be uniform throughout the 

 large number of lines, the following method of culture was found 

 necessary : 



As a culture medium, Horlick's malted milk was employed, 

 following the example of Miss Peebles ('12), one-sixteenth of 1 

 per cent being found the most favorable proportion. This was 

 made fresh each day, with boiling water. The animals were 

 changed every other day. The chief difficulty in making the 

 conditions uniform throughout all the lines is as follows: A 

 nimiber of diverse bacteria are found in the cultures, falling into 

 them at the time of changing, or reaching them in other ways. 

 Some of these multiply strongly in certain of the slide cultures 

 while others get a better foothold in others. The effect of the 

 diverse bacteria on the rate of reproduction differs greatly; as a 

 result therefore some of our lines of Paramecium multiply rapidly, 

 others slowly, even though there is no intrinsic differentiation 

 among them. Now, in transferring with a capillary pipette a 

 single individual to a new drop, as is done at the time of changing 

 the animals, inevitably a certain amount of the bacterial culture 

 is transferred with them, serving to infect the new drop. Thus' 

 one line will be accompanied always by the bacterium x, causing 

 rapid multiplication; another by the bacterium y, causing slow 

 multiplication. The results simulate those of inherited differen- 

 tiation in the fission rate. 



Experience showed that this difficulty is obviated by the fol- 

 lowing method of procedure, which was adopted for the present 

 experiment on August 28: 



The new fluid (1/16 pisr cent malted milk) was made sterile 

 by boiling. It was then infected with bacteria from a mass 

 culture of the race E, in which the animals were flourishing 

 strongly. This was done by filtering (through two thicknesses 

 of filter paper) a quantity of the fluid from this culture (in order 

 to remove the Paramecia). I added four pipettes full of this 

 filtered fluid to 100 cc. of the fresh culture fluid. 



