CHEMICAL STIMULATION OF OVULE DEVELOPMENT AND ITS POSSIBLE RELATION TO PARTHENOGENESIS

JUST AS the production of polyploid plants by the colchicine technique has proven of value in plant breeding, so the discovery of a method for producing haploids at will would also be a useful tool. Since the haploids which occur spontaneously'in nature probably arise through reduced parthenogenesis, this would seem to be the most likely means of attaining the desired result. If this procedure could be carried out by artificial methods, such as chemical treatment, a new way would be available for establishing pure lines. Thus a 2n plant could be treated to yield a In plant bearing only maternal characteristics. The chromosome number of this plant could then be doubled by colchicine treatment, producing a plant which would breed true. With this in mind, a study of the effect of various chemicals in stimulating fruit, seed and egg-cell development was made in order to survey the possibilities of producing haploid plants by artificial treatment. Many inorganic and organic chemicals, including plant hormones such as auxins,2 traumatic acid, biotin and pantothenic acid, were injected into the ovaries of plants to observe their effect. Although pseudoembryos were obtained in Datura after auxin treatment, no true embryos resulted from any chemical tested. As a result of observations made in these experiments, combined with information obtained by a survey of the literature on parthenogenesis, haploidy, pseudogamy and related subjects, a working hypothesis of fruit and seed development has been suggested. MATERIALS AND METHODS.-A 4n strain of fieldgrown Melandrium dioicum and both 2n and 4n strains of Datura stramonium were used as test plants. Datura was chosen because detailed information concerning its characteristics is available. Melandrium was chosen because it is dioecious, and because of the nature of the ovary which, with its central placentation, surrounded by a large empty space, greatly facilitates injection with various solutions or emulsions. Flowers to be treated were cut just above the ovary when the buds were about to open. In this way the styles were removed, and in the case of Datura, the anthers also, thus preventing stray pollination. Immediately thereafter the ovaries were injected, by means of a hypodermic syringe, with the various chemicals. Since the Datura ovaries are quite solid and divided into four locules, injections were made through the side of the ovary wall in two to four places. In Melandrium, injections in the side of the ovary proved somewhat injurious to the fruit, frequently causing fungal infection. Infections were avoided by making injections through the top of the 1 Received for publication April 28, 1941. 2 Any substance causing cell elongation in the Avena test is by definition [66] an auxin. ovary. One hole was made to allow air to escape, and another for the inj ection, thus filling the entire space around the ovules with solution. The chemicals were tested in a wide range of concentrations. For each concentration a minimum of ten ovaries was treated. Only the auxins naphthaleneacetic acid and indolebutyric acid prevented abscission of the fruit. Since fruit retention is essential to a further study of the effect of treatment, it was the usual practice to inject a solution of auxin in combination with any other substance to be tested. In order to increase the solubility of the auxins they were converted to their ammonium salts by adding a few drops of dilute ammonium hydroxide until they dissolved. A stock solution of one per cent in distilled water was diluted to give solutions of the desired strength. Auxins were also applied in the lanolin emulsion made with morpholine which has been used successfully as a vehicle for colchicine [62]. A 0.1 per cent auxin-lanolin-emulsion was obtained by adding one volume of stock auxin solution to nine volumes of plain emulsion. The emulsion was applied internally as well as externally by using a regular hypodermic syringe. OBSERVATIONS.-Auxin treatment of Melandrium. -When the styles of Melandrium are removed as outlined above, but not injected with auxins, the ovaries abscise within a few days. Naphthaleneacetic acid, indolebutyric acid, and indoleacetic acid separately and in combinations were injected into Melandrium ovaries in concentrations ranging from 0.000001 per cent to 1.0 per cent. Injection of 0.1 per cent naphthaleneacetic acid into the ovaries gave by far the best results, at least 90 per cent of the ovaries of Melandrium so treated yielding parthenocarpic fruits. Lower concentrations resulted in a very high percentage of abscissions, whereas higher concentrations damaged the ovaries. The size of such parthenocarpic fruits was approximately the same as that of fruits obtained by normal pollination. Indolebutyric acid gave fairly good results but not as good as naphthaleneacetic acid. Indoleacetic acid failed to give satisfactory results, perhaps because it is less stable than naphthaleneacetic acid. Experiments were undertaken to find out whether the auxins could be applied externally, thus preventing injuries from the injections. Emulsions containing auxins (usually naphthaleneacetic acid) were therefore applied, (a) on the peduncle, just below the flower, (b) on the outside of the ovary, (c) in the ovary by injection. The average outside dimensions of fruits resulting from these treatments were nearly the same (fig. 1; table 1). However, when they were opened, great differences were found (fig. 1; table 1). Whereas in ovaries that had been treated externally, the placenta and the ovules had hardly en-