Cytokinin effects on pollen embryogenesis in cultured anthers of Hyoscyamus niger

Abstract

Addition of the cytokinins, benzylaminopurine, N6-Δ-(2-isopentenyl)-adenine, kinetin, zeatin, and zeatin riboside to a basal medium containing mineral salts and sucrose induced characteristic changes in pollen embryogenesis in cultured anthers and anther segments of Hyoscyamus niger. In anthers cultured in media containing 0.01–10.0 mg/L of each cytokinin, there was a progressive delay in the appearance of plantlets outside the anther locule and in the morphology of plantlets formed. Among the cytokinins tested, only zeatin riboside promoted anther efficiency; however, all compounds reduced pollen efficiency by as much as 40–60% of the control even in the lowest concentration tested. The effects of cytokinins were particularly noticeable in the failure of pollen grains to form embryoids by the division of the generative cell and in the decrease in the number of embryoids formed by the division of both generative and vegetative cells. At the morphogenetic level, embryoids formed in media containing cytokinins displayed various abnormalities such as precocious elongation of the root apex, cotyledon expansion, and callus formation on the cotyledons, hypocotyl, and root. During their subsequent growth, calluses induced on embryoids by benzylaminopurine, N6-Δ-(2-isopentenyl)-adenine, zeatin, and zeatin riboside formed somatic embryos that recapitulated stages of zygotic embryogenesis. Globular and heart-shaped embryoids, which did not develop further, were frequently observed on kinetin-induced pollen callus; the callus also regenerated leafy shoots by organogenesis. Addition of adenine to the medium did not have any effect on pollen embryogenesis in cultured anthers of H. niger.