The Isolation And Identification Of Vegetative Storage Protein In Ginkgo Biloba L. And Its Characteristics Research

In this paper, the anatomic study on ultrastructure of phloem parenchyma of Ginkgo biloba L. in different periods was researched as the breakthrough, connected with histochemistry analysis and biochemistry analysis, the technology of immunocytochemistry localization with enzyme and colloidal gold was applied to do exactly localization for vegetative storage proteins of Ginkgo biloba L.. The components of vegetative storage proteins in Ginkgo biloba L., whose separation and purification and whose property of glycoprotein were analyzed by the technology of SDS-PAGE and different dye methods, and the main components of vegetative storage protein of Ginkgo biloba L. were determined. The correlation between the seasonal dynamic change rule of vegetative storage protein and the growth and development of Ginkgo biloba L. were clarified. The basic rules of formation, accumulation and degradation on vegetative storage protein in Ginkgo biloba L. were discovered. The above studies would have important theoretical value and practical significance for improving the utilization efficiency of nitrogen, guiding the rational application of fertilization and cultivating fine variety with high utilization efficiency of nitrogen by transgenic technology. Main conclusions were as follows:The vegetative storage proteins were greatly distributed in the phloem parenchyma cell, the phloem radiation ray cell, the xylem ray cell, and the cell of between secondary phloem and primary phloem of branches and roots in Ginkgo biloba L., whose number was the most in the phloem parenchyma cell, which were synthesized in cytoplasm and endoplasmic reticulum and Golgi apparatus were founded in the course. The vegetative storage proteins in vacuole were in the form of irregular block, flocculent structure, granule structure and thin sand.The vegetative storage protein of Ginkgo biloba L. mainly located in phloem and xylem of branches, whose content in one-year branches was higher than that in two-year branches, whose content in phloem was higher than that in xylem. The dynamic change rule of vegetative storage protein might be divided into two phase of accumulation and degradation. With the new shoots growth and new leaves formation in May, vegetative storage proteins were accumulated in new shoots and whose number increased greatly, and whose content amounted to the most in December. In the whole winter, the content of vegetative storage protein did not change. When the buds began germination, vegetative storage proteins were rapidly degraded, transformed and utilized to meet the demand of the growth of new shoots, and which were degraded fully on the 14th in April.The vegetative storage proteins of Ginkgo biloba L. could be divided into three kinds of proteins with high molecular weight, middle molecular weight and low molecular weight. One protein with the same molecular weight might have different isoelectric point. Vegetative storage proteins of Ginkgo biloba L. were determined, whose molecular weights and isoelectric points were respectively 36.4 and 5.56, 36.3 and 4.57, 36.1 and 5.35, 36.4 and 5.49, 36.2 and 6.46, 36.2 and 5.02, 36 and 6.33, 36 and 6.74, 36.1 and 4.62, 36.4 and 5.19, 36 and 6.2, 36 and 6.43, 32.3 and 5.15, 32.4 and 4.66, 32.1 and 6.6, 32.1 and 6.3, 32.2 and 4.74, 32 and 5.97, 32.1 and 6.45, 32 and 5.75. The vegetative storage protein of Ginkgo biloba L. had typical glycoprotein property.The antigens of vegetative storage protein of 32kDa and 36kDa were separated and prepared by SDS-polyacrylamide gel electrophoresis, and the new Zealand rabbits were immune for four times with 32kDa and 36kDa antigens and whose blood was collected to extract antiserum. The antibodies of two proteins were tested by the two-dimensional agar immunodiffusion test, which suggested that antibodies of vegetative storage proteins of 32kDa and 36kDa were achieved successfully. The trial of the immunogold electron-microscopy localization and immunoenzyme optical-microscopy localization also illustrated that proteins of 32kDa and 36kDa were the main vegetative storage proteins of Ginkgo biloba L.