392 
MABEL MARY BROWN 
The capsules were opened by means of needles sterilized in the flame of a 
Bunsen burner, and the spores were then scattered on the surface of the 
culture fluid. In order to make the sowing more uniform, the sporophyte 
was held a short distance above the medium and the capsule was tapped with 
a needle. If the spores are allowed to germinate in groups, the protonemata 
become interlaced and can not well be separated. The cultures were 
examined daily under a binocular microscope in order to note the time re- 
quired for spore germination, and the development of rhizoids and branches. 
The removal of the individual spores from the petri dishes to soil was 
attended with some difficulties. The very young protonemata can not 
be caught up on a needle, so it was necessary to have recourse to the use of a 
pipette and spore dilutions. Great care must be taken to make certain 
that but one gametophyte is transferred to each pot. A careful examination 
of the contents of the pipette, and of the surface of the soil after these 
contents have been deposited upon it, should be made. The procedure de- 
scribed above was used for the first lot of cultures made. Th'en it was found 
that if the gametophytes are allowed to grow until the protonemata are at 
least seven or eight cells in length, it is comparatively easy to pick them from 
the culture fluids, under the binocular microscope, by means of a needle. 
Three kinds of soil were used for the single spore cultures: a mixture of 
clay loam and sand ; a mixture of leaf mould and sand ; and a soil containing 
a large amount of wood ashes. The last named soil was obtained from a 
spot where Funaria hygrometrica had been growing. The soils were placed 
in two- and three-inch pots, soaked with Marchal's solution, and sterilized 
in the autoclave at from ten to fifteen pounds pressure for at least three 
hours. After cooling, the soils were ready to receive the young protonemata. 
Four hundred twenty-five single spore cultures were made at different 
times from the last of November, 1916, until the last of March, 1917. 
The cultures were placed in a Wardian case in the greenhouse, where the 
atmosphere was kept very humid and where the temperature ranged from 
about 70° in the winter to as high as 112° F. at times during the summer. 
The pots were set in earthenware plates in which water was kept so that 
the cultures received water only from below. In this way the danger of 
contamination by foreign spores and other organisms was minimized. 
The plants grew well and by the last of June, 191 7, all the pots were 
covered with gametophores. These cultures have been kept for about two 
years and all have remained pure, no moss of other species appearing in any 
of the cultures. 
