nmm sP

TECHNIQUE 41
be undesirable. Weak agar media made with tap-water, dilute Knop solution
or 1 per cent, malt are.employed when it is desired to isolate individual sporidia
in series from single promycelia of Ustilago species.
Sterihzation of chlamydospores can be effected by soaking them for 24 hours
or longer in 1 per cent, copper sulphate solution (Stakman et al., 1929).
MEDIA FOR GROWTH IN CULTURE
Smuts will grow on many standard media used for fungi, but slight changes
in composition alter the appearance of the colony. For tte comparison of
several monosporidial lines cultures are made in dupUcate or triplicate on one
batch of the medium, the same quantity of agar being poured into each dish.
Potato glucose and potato sucrose agars have been widely used for the study of
gametophytic characters (see p. 30).
In descriptive work shades of colour are matched by standard plates such as
Ridgway's. Photographs are useful to record features not easily described and
cultures are usually in good condition for this after about 40 days. To avoid
disturbing reflections liffthe circle of agar bodily from the dish and place it on
a sheet of cardboard.
To maintain stock cultures in an active condition they should be transferred
every four weeks. Potato glucose agar (1 per cent.) and a modified form of
Czapek agar have been used for U. maydis. The second medium has the merit
of lowering the rate of mutation (see p. 37). Meat extract agar (1 per cent.) was
used for stock cultures of the oat smuts by Sampson & Western (1938).
Smuts attacking grasses have not yet been widely cultured. Fischer (1940)
found that U. striiformis wiU tolerate a relatively high concentration and makes
good growth on agar containing 8 per cent, dextrose, 4 per cent, malt extract,
and 1 per cent, peptone.
No infallible technique can be given for inducing the development of chlamydospores
in artificial culture (see p. 25). Species of Entyloma form them readily
in a few weeks on potato dextrose agar. Cultures may be started by fastening
portions of fresh, infected leaves to the hds of .Petri dishes, allowing the sporidia
to fall, as discharged, on the surface of clear filtered agar, and selecting those
colonies that originate in the larger, allantoid, binucleate sporidia (see p. 22).
Chlamydospore formation is long delayed in some species, occurring in Tilletia
caries only after three months on oatmeal and Leonian agars (BuUer, 1933).
Schmitt (1940) tried many media to induce the formation of chlamydospores in
U. maydis without success.
PREPARATION OF MONOSPORIDIAL CULTURES
For genetical analysis it is necessary to isolate a series of single sporidia in a
particular order from the promycelium of one chlamydospore. Dickinson (1926)
described an apparatus designed to move a finely pointed glass needle over a
small field in three planes, and to drag small cells, from 30 to 20 /n down to the
Hmits of microscopic vision, to a chosen part of the field while under observation.
Haima (1928) constructed a similar isolator from parts of apparatus commonly
used in a laboratory and employed it for the preparation of monosporidial
cultures of Ustilago maydis and Sphacelotheca reiliana. A single chlamydospore