Methods for non-stigmatic pollination in Trifolium repens (Papilionaceae): seed set with self- and cross-pollinations in vitro.

The stigma and style of Trifolium repens L. was successfully by-passed, and viable seed was obtained by placing pollen on a perforation made in either the ovary wall or in the base of the style. The application of pollen to the stump of the style also succeeded in producing viable seed. The presence of the calyx and corolla facilitated seed set in cultured florets. An incision made along the entire length of the ovary to permit pollen entry resulted in necrosis of the floret. Perforation of the ovary did not affect the yield of seeds obtained from stigmatic pollinations. Growth of pollen tubes was observed on wounded tissues and also within the style and cavity of the ovary. A mean yield of 20 embryos was obtained per 100 florets in non-stigmatic pollinations, whereas with stigmatic pollinations in vitro the yield was 148. Nonstigmatic pollinations also gave seeds when T. repens was self pollinated, which showed that the gametophytic system of self-incompatibility (SI) could be by-passed. With one genotype, self-pollination via the stigma failed, whereas embryos were obtained via non-stigmatic pollinations.