278 ARTIFICIAL PARTHENOGENESIS AND FERTILIZATION 
connection with the experiments, showing beyond a doubt that the 
female plants treated were absolutely sterile. 
Loeb has shown that, when unfertilized eggs of the sea-urchin are 
placed for one and one-half to two minutes in a mixture of 50 c¢.c. of 
sea-water+3 c.c. of 0.1 m acetic, butyric, or other fatty acid and then 
transferred to normal sea-water, a fertilization membrane is formed. 
This method was applied to unfertilized Fucus eggs. In experimenting 
with the eggs those used at any one time were always divided into three 
lots. One lot was used as a control, another was fertilized, and the 
third was treated with the solution. If a single egg in the control 
formed a cell-wall, which seldom happened, the three lots were dis- 
carded. In case the eggs were treated with acetic or butyric acid, 
as above described, a large number of them formed in about ten 
minutes a membrane or cell-wall which was exactly similar to the one 
formed about normally fertilized eggs. By plasmolyzing the eggs the 
membrane is readily seen. Eggs not treated with a solution or not 
ertilized undergo cytolysis and degenerate. In any case many of the 
eggs failed to develop, but about one-fourth as many formed mem- 
branes under the influence of the solutions as were formed about ferti- 
lized eggs. After the formation of the membranes if the eggs are 
placed in hypertonic sea-water, 8 ¢.c. to 10 ¢.c. of 2.5m NaCl or KCl 
+50 c.c. sea-water, for 30 minutes and are then brought back into 
normal sea-water, development continues. Nearly all of the eggs 
which have formed a membrane become pear-shaped, showing a 
rhizoidal papilla, and by next morning have cleaved. The rhizoidal 
cell is cut off and one or more cleavages have taken place in the other 
portion of the sporeling. If the cultures are properly aerated, spore- 
lings develop resembling in every respect those grown from fertilized 
eggs. In place of sea-water containing a fatty acid, solutions of 
various other cytolytic substances were used, but none stimulated 
membrane formation or development as well as the acids. 
With regard to the first formation of the cell-wall over the surface 
of isolated masses of plant protoplasm, it is usually attributed to a 
process of secretion by the outer layer. That the process is a rapid 
one is shown by the fact that in Fucus eggs a cell-wall is formed in ten 
minutes after the entrance of the sperm. Cell-wall formation may also 
be artificially induced, as shown above, by various substances. In 
some cases a cell-wall may appear under certain conditions on the sur- 
face of plasmolyzed protoplasts in fifteen minutes, as has been shown 
by Klebs, Palla, and others, while in other cases hours are required 
