434 LORANDE LOSS WOODRUFF AND RH. ERDMANN 



in a general way by periodically preserving specimens left over 

 at the daily isolations. 



The culture was started and conducted for two months at the 

 Biological Laboratory of Williams College, and since has been 

 carried on at the Biological Laboratories of Yale University, 

 except during a part of each summer when it was transferred 

 to the Marine Biological Laboratory at Woods Hole, Mass. 



It is under these conditions then that this culture (I) has been 

 successfully maintained without loss of vigor for over 4500 gen- 

 erations — all the experiments which have been conducted on 

 animals from this race having been made on sub-cultures started 

 from the animals left over from the lines of this culture at the 

 daily isolations. 



B. METHOD OF CONDUTCTION OF SUB-CULTURE IE 



For the intensive study of the daily cytological changes of 

 this race of Paramaecium aureha, on October 27, 1913, six ani- 

 mals left over from Line la of the main culture at the 4020th 

 generation were isolated to start a new sub-culture, designated 

 IE, of six lines. This sub-culture was subjected to practically 

 constant environmental conditions. The culture mediimi was 

 the 0.025 per cent beef extract which Woodruff and Baitsell 

 ('11 a) found to be a most favorable medium for this race of 

 Paramaecium. The sub-culture was kept in a thermostat set 

 for 26°C., and such variations (about 1°C.) from this as occurred 

 were well within the optimum zone for the animals of this race 

 as determined by Woodruff and Baitsell ('11 c). 



An animal was isolated every day from each line of this sub- 

 culture from its initiation to April 27, 1914 — a period of six 

 months. The isolated animal from each line was placed in fresh 

 culture medium, while one or more of the remaining animals was 

 preserved for study. Thus for half-a-year permanent prepara- 

 tions were made from day to day of sister cells, the exact ancestry 

 of which was known in every case. It is important to emphasize 

 the fact that by this method only, i.e., the study of practically each 

 cell generation, could the sequence of nuclear changes which we 

 describe be determined. This became evident very soon as the 



