An indirect method to assay for mitotic centers in sand dollar (Dendraster excentricus) eggs.

It is possible consistently to induce sea urchin and sand dollar eggs to cleave directly from one cell into four cells. This is done by exposing the fertilized eggs to benzimidazole for 20 to 30 min beginning about early metaphase. The mitotic apparatus regresses, the cells do not cleave, and shortly after they are returned to normal sea water an early-prophase-appearing nucleus is present in each cell. Each cell then organizes a tetrapolar tetrahedral mitotic apparatus de novo, instead of transforming a bipolar mitotic apparatus into a tetrapolar figure, and cleaves one-to-four. In another type of experiment, it appears that sand dollar eggs exposed to mercaptoethanol during the first period of mitotic center duplication have only half as many centers by first cleavage metaphase as the normal controls. This is consistent with an earlier report by Mazia et al (1960). Using this same experimental technique, it was demonstrated that benzimidazole, on the contrary, does not interfere with mitotic center duplication in sand dollar eggs. A labeling experiment demonstrated that benzimidazole does not interfere markedly with the normal pattern of incorporation of C(14)-thymidine into the DNA of sea urchin eggs. The data reported here suggest that judicious treatment of sand dollar eggs (and probably sea urchin eggs, too) with benzimidazole can induce the eggs to cleave into as many cells as there were mitotic centers sometime earlier, for example at early metaphase of the first cleavage division. This provides a very useful tool for studies on the process of mitotic center duplication.