| This dissertation focuses on the mechanism of somatic embryogenesis in established embryogenic suspension cultures. A detailed light microscope study of the cellular characteristics, regenerative potential and mode of somatic embryogenesis of all culture components. The results show that established embryogenic suspension cultures of daylily, probably all plants, are already determined in the primary culture stage, well before a culture is perpetuated through subculture. The conditions of maintenance culture perpetuate a determined state in the form of a proliferative collection of initials limited in their development but undergoing what might be termed a ‘forced polyembryony’. Evidence showing that the role of auxin in maintenance culture is to stimulate multiplication of determined but immature embryos and thus its removal is permissive for regeneration.; Experiments establish that the regeneration phase medium modifications and additions beyond hormone removal/reduction that are required to achieve/optimize daylily somatic embryo development. The common empirical practice of utilizing semisolid medium to achieve somatic embryo regeneration is a permissive modification. By re-examining the process using light and scanning electron microscopy, an embryogenic developmental process occurs when plantlets are regenerated from suspension material entirely in liquid medium via structures referred to in this laboratory as pre-shoots.; The earliest events in the formation of daylily suspensions initiated with shoot tips (used exclusively in these studies) shows, in accord with the characterization put forth here, that determination occurs in the primary phase of culture. Primary responses are morphogenetically competent and derive from parenchymal cells (probably from single cells) associated with the vasculature of leaf bases unavoidably retained on the explant (surrounding the apex) rather than, as suspected, from meristematic cells of the apex. Even as the formation of high quality mature embryos will not occur at any time in a medium containing the high levels of growth regulators levels found in the induction/maintenance medium, some of the early responses compare favorably with globular stage somatic embryos. Ultimately, nodular structures develop that later proliferate as nodular masses. The suspension derives as what may be viewed as buds from this relatively short lived primary response. Discussions are presented to suggest that the type of primary response seen in daylily is extremely common and that it is not true callus. (Abstract shortened by UMI.)... |
