250 
METHODS IN PLANT HISTOLOGY 
The various stages of the female gametophyte and embryo are 
not hard to stain; but the walls throughout the development of the 
male gametophyte are very thin and extremely hard to stain. 
Safranin and light green is a good combination. Light green in clove 
oil may prove more satisfactory than the alcoholic solution. 
Isoetes. —This peculiar genus is widely distributed and 16 of 
its 64 species occur within the United States. It looks so much like 
a sedge that it is easily overlooked, even when rather abundant. 
As a genus, it is hydrophytic, growing in wet places or even under 
Water. A recent monograph by Dr. Norma Pfeiffer not only gives 
an ecological, morphological, and taxonomic account (with keys in 
English), but gives hundreds of stations, a feature which will enable 
many to find material. 
Vegetative structure— The short, thick stem, even in old plants, 
cuts easily in paraffin. Fix in formalin alcohol and stain in safranin 
and light green. Sporelings with stems about 2 mm. in diameter and 
young plants with stems up to 5 mm. in diameter are best for a 
study of the peculiar vascular system of this plant. These young 
stages fix well in chromo-acetic acid and are not hard to cut. 
Sporangia. —All the sporangia of the plant may be said to con¬ 
stitute a single strobilus of the Selago type. Both longitudinal and 
transverse sections should be cut. The stem is so short that, in 
a plant of medium size, a longitudinal section may include the stem, 
the sporangium, and the sporophyll, up to the top of the ligule. 
Such sections, 10 to 15 n, or even 20 m in thickness, are best for 
demonstration. Transverse sections through the whole cluster of 
sporophylls show the arrangement of megasporophylls and micro- 
sporophylls and also the relations of the sporangia to sporophylls. 
The gametophytes. —The spores are shed in the uninucleate stage, 
and consequently it is not so easy to find the germination as in the 
case of Selaginella. When the large megasporangium begins to 
decay, let the megaspores dry naturally. They retain their power 
of germination for a year at least. Simply wet them with tap water 
and the earlier stages are easily secured, quite clean and ready for 
cutting. There must be soil in the dish for later stages. Try a 
similar method for microspores. Also, look at the top of the stem 
of old plants for stages developing naturally. The cell walls of the 
male gametophyte, as in the case of Selaginella, are rather hard to 
differentiate. Use anilin blue or light green. 
