SUMMARY

The focus of this research is to gain insight into the role of mechanical stress in the development of plant somatic embryos in terms of synchronized development and yield of mature embryos on solid and in liquid culture medium. Practical implementation of somatic embryogenesis for large-scale clonal propagation of plants and agricultural goods is hampered due to non-synchronous development of embryos resulting in low yields of plants. This doctoral research hypothesizes that dispersion of clusters of proembryogenic masses (PEMs) provides more equal growth conditions to all somatic embryos and thus aids in more synchronization of embryo growth and development. The research includes the design of a temporary immersion bioreactor, the design of dispersion system, growing somatic embryos of Norway spruce (Picea abies) as control and experiment on solid medium and in temporary immersion bioreactor, dispersion of the clusters of PEMs, image analysis of the dispersed PEMs to get the particle size distribution in order to determine the effect of dispersion on embryo growth and development, image analysis of mature embryos to get the size distribution to evaluate the level of synchronization of embryo development, and the design of automatic dispersion system to impose periodic, intermittent stresses on PEMs during proliferation stage of embryo development. Results indicate that dispersion of aggregates of PEMs of Norway spruce has a positive effect on both the rate of proliferation of PEMs and the subsequent development of mature somatic embryos both on solid and in liquid medium. To investigate in further details to understand the association between development of mature embryos and nutrient uptake by cells and tissues, a nutrient diffusion model is developed using the volume averaging technique. It estimates the concentration distributions of nutrients, e.g. sugars, in the PEMs clusters on solid medium over the period of sub-culture. Michaelis and Menten enzyme kinetics is used in the model for the uptake of nutrients by the cells and tissues. Enzymatic assaying of soluble sugars is performed to determine the concentration of sugars (glucose, fructose, and sucrose) at different locations in the tissue clusters. Experimental results are compared with the model outputs. A significant match between the predicted and experimental outputs is observed.