Self -Fertilization Induced by Artificial Means. 137 



of the rising sun. The rough handling incidental to removal and 

 isolation appears to cause Ciona to retain its eggs for several 

 days. The individuals to be used were isolated, as a rule, from 

 24 to 48 hours, and in most cases were rinsed in fresh water before 

 opening. It was not found necessary to boil the water; for check 

 experiments "showed that eggs left to themselves were never fer- 

 tilized by stray spermatozoa in the sea-water. Since Ciona de- 

 posits its eggs only in the very early morning, the chances are 

 very slight that functionally active spermatozoa would be present 

 in the sea-water in the late morning and in the afternoon when 

 the experiments were carried out. 



The eggs of Ciona are surrounded by a rather thick membrane. 

 Standing out like broad spikes over the surface of the membrane, 

 and forming a beautiful aureole around the egg, are the trans- 

 parent follicle cells, each with a shining drop in its outer end. 



A'number of preliminary experiments confirmed Castle's con- 

 clusion that self-fertilization is rarely possible in Ciona intes- 

 tinalis. The evidence, however, on which Castle based this con- 

 clusion is not altogether satisfactory, since he records many cases 

 in which self-fertilization occurred. Instances are cited in which 

 isolated individuals gave 90, 25, 16, 5, 4, o per cent, of self- 

 fertilized eggs. Castle supposes that, in the first of these cases 

 at least, the spermatozoa of one day fertilized the eggs of the 

 next, but it has not been shown that the spermatozoa have this 

 power if left so long in sea-water. The same individuals that 

 had been used for these isolation experiments were killed (after 

 being washed in 90 per cent, alcohol), and the eggs and sperm 

 of each taken out and mixed together. The results gave 50, 4, 

 I, Ys, o per cent, of self- fertilized eggs. The same exj^rriment 

 repeated with fresh individuals gave 50, 12^, 10, 5, 2, o per cent, 

 of self-fertilized eggs. From these figures it is clear that in some 

 cases a considerable amount of self-fertilization occurred, unless 

 there was some source of error in the experiment. In fact. Castle 

 believes that in those cases where a large number of eggs were 

 fertilized there was some contamination. My own results with 

 Ciona have never given so large a percentage of self-fertilized 

 eggs, and I am inclined to attribute this result in part to the 



