EMBRYOLOGY IN NORWAY SPRUCE (PICEA-ABIES) - AN ANALYSIS OF THE COMPOSITION OF SEED STORAGE PROTEINS AND DEPOSITION OF STORAGE RESERVES DURING SEED DEVELOPMENT AND SOMATIC EMBRYOGENESIS

Seed cones were collected from open-pollinated trees of Norway spruce (Picea abies) in a seed orchard from Pollination until maturation of the seeds. Immature embryos were isolated for embryogenic tissue cultures that were maintained either on solidified medium or as liquid cultures. By transferring young somatic embryos to medium containing 90 mM sucrose and 7.6 muM ABA growth continued to mature embryos that accumulated storage reserves in both the hypocotyl-shoot axis and the cotyledons. Both zygotic and somatic embryos at different developmental stages were processed for microscopy as were the megagametophytes. Total protein was extracted from the seed material at intervals during development and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These analyses revealed that storage protein started to accumulate in the megagametophytes at the time when embryos were growing into the gametophytic tissue, while it occurred a few weeks later in the embryos at rapid embryo growth and organ differentiation. Lipid bodies also became abundant in the mature plant material. Although plastids with prominent starch grains were very frequent in both megagametophytes and embryos during development they were not observed in the desiccated tissues. Zygotic and somatic embryos displayed a similar developmental pattern. By sequential salt-extraction and dilution two fractions highly enriched in storage protein were obtained. One fraction (G-1), requiring higher salt concentration for protein solubilization, was dominated by a protein migrating to around M(r) 55 000-60 000 when separated under non-reduced condition. After exposure to reducing agent this protein was replaced by two new ones with M(r) 33 000 and 22 000 giving evidence of disulfide bonded polypeptides. The other fraction (G-2), was dominated by polypeptides around M(r) 42 000 and low molecular mass polypeptides (< 14 000).