Histological events leading to somatic embryo formation in cultured petioles of alfalfa

Summary

An optimum 10-day exposure of petioles of alfalfa [Medicago sativa ssp.falcata (L.) Arcangeli] to 2,4-dichlorophenoxyacetic acid or 2,4,5-trichlorophenoxyacetic acid results in the semisynchronous production of somatic embryos starting about 4 days after transfer to a non-auxin-containing medium. The timing of cell division induction in the petiole explants was found to vary depending on the petiole tissue type. Cells adjacent to the vascular bundles divide first at about 48 h after exposure to auxin, closely followed by those of the inner parenchyma, whereas most of the cells of the subepidermal and epidermal layers start to divide later, between 72 and 120 h. Two different sources of callus were also evident. Cells adjacent to the vascular bundles and the inner parenchyma cells were the primary source of callus when a short, 2-day (non-embryo-producing) exposure to auxin was used. In contrast, the subepidermal and epidermal cells were the primary source of callus tissue when a longer, 10-day (embryo producing) exposure was used. It is concluded that the source of somatic embryos is primarily the daughter cells of the subepidermal or epidermal tissue or both.