Nuclear Transplantation in Amoebae. I. - Journal of Cell Science

Lorch and Danielli—Nuclear Transplantation in Amoebae. I 447
by means of a mouth pipette and distributed in solid watch-glasses containing
2 ml. of a 'food culture' of ciliates or flagellates, or a mixture of both in the
following modification of Chalkley's (1930) medium:
Stock solution
NaCl
NaHCO 3 .
KC1
Na 2 HPO 4 .i2H a O
Water (glass-distilled)
160 gm.
08 gm
0-4 gm.
0'2 gm.
1000 ml.
To make 1,000 ml. of culture medium add 5 ml. of stock solution to 995 ml.
of glass-distilled water. The pH is about 7-5.
The success of the experiment depends on the food culture. The most
suitable food organisms were found to be the flagellate Chilomonas paramecium
and the ciliate Colpidium sp.
The food culture in the watch-glasses was sucked out and replaced by fresh
culture every 4-7 days, depending on the rate of multiplication of the amoebae
and on bacterial growth. Amoebae do not thrive when dense bacterial growth
is present.
When the number of amoebae in a solid watch-glass exceeded 30 (i.e. after
10-14 days) the contents were transferred to a Petri dish containing food
culture. Here the amoebae may be left for 3-4 weeks, the food organisms being
replenished if necessary. After this time the culture was usually well established
and ready for transfer to the standard culture dish. This is a shallow
pyrex dish, 10 cm. internal diameter, filled with 100 ml. of Chalkley's medium
to which 4 boiled wheat grains were added. The culture dishes were kept
covered, in dim light, at room temperature (18-23 0 C). Every 4 weeks the
cultures were examined under a dissecting microscope, most of the fluid and
2 of the wheat grains were sucked out, and fresh Chalkley's medium and
2 boiled wheat grains were added. Every 2 or 3 months the cultures were
transferred to clean dishes. There is considerable variation in the state of
amoeba cultures even when established under relatively standard conditions;
therefore the treatment of the mass cultures was according to need rather
than rigidly standardized. In some dishes amoebae multiplied rapidly. In these
cases the food organisms have to be replenished from time to time. In other
dishes there appeared dense bacterial growth which led to a rapid increase in
the ciliate population and a decline in the number of amoebae. The latter
became sluggish and were attacked and eaten by the 'food' organisms. The
cultures were always kept free from rotifers, worms and Crustacea. Algae do
not appear to be harmful but were excluded because in their presence it is
difficult to see the amoebae clearly. Various moulds were noted on the wheat
grains. They cannot be avoided in non-sterile cultures and are certainly
not harmful. The mould hyphae are often densely covered with amoebae
whose extended pseudopods form a network enmeshing the ciliates and
flagellates which feed on the bacteria accumulating round the decaying
wheat.