Comparative Studies On Dissection Structure And Localization Of Key Photosynthetic Enzyme Among Two Ephameral Plants,C3 And C4 Plant

Ephemeral plant is a special plant species which is able to make full use of melting snow water and spring rainwater, as well as the illumination and heat resources,to start and soon finish a life cycle within 60-90d before hot summer season is coming. They distribute only in the north of Tianshan mountain, Jungar Basin and the surrounding arid desert area of Xinjiang province, China, and therefore play important role in reducing sand storm, wind-breaking and sand-stabilizing, protection and improving ecological environment. Moreover, there have a large amount of species which are valuable in feeding, medicinal and decorative purpose, so ephemeral plants consist of the biological resource pool with unique and very high potential value for utilization. Recently, the characteristics of high photosynthesis efficiency, short life cycle, fast growth and development of the ephemeral plants have already become a hot research field for biologists.We choose two ephemeral species and two Carbon three plant species as materials: Eremopyrum orientale, Lepidium apetalum, Brassica napus and Wheat cultivar. Several experiments and research work on microstructure, ultrastructure and immunogold localization showed the characters of the four plants on anatomical structure, photosynthetic cellular organelle and the localization of photosynthetic key enzyme, then compared the differences between ephemeral plants and Carbon three plants, finally discussed the pattern of photosynthetic carbon metabolism of ephemeral plants.The cellular structure of leaves was observed under light microscope by using paraffin section technique. The results showed that the tracheary element in vein of Eremopyrum orientale and Lepidium apetalum are bigger than another two Carbon three plant species, and this wide tracheary element is available for rapid absorption and transportation water in short time.The major water-storing parenchymatous cells which are making for keeping water can be observed in mesophyll of ephemeral plant. There are lots of small chloroplasts in bundle sheath cells(BSC) around vascular bundle. For these chloroplasts distributing along the edge of BSC, it looks like a wreath, so called"Kranz type"which is the typical structure of Carbon four plant. Above-mentioned special physiological structure of ephemeral plant is formed for adapting to the environments in arid and wind-striking area in long period of evolution.The anatomic structure was observed under transmission electron microscope (TEM) by using resin ultra-thin cross-section technique. The results of ultrastructure indicated that the cellular organelle in BSC of two ephemeral plant species were more abundant than that of another two Carbon three plants. The grana lamellae in chloroplast in BSC of ephemeral plants are underdevelopment. And a lot of starch grains are presented in BSC of ephemeral plants.Because Rubisco is a key enzyme in photosynthesis, immunolocalization of Rubisco in Carbon three plants and two ephemerals was determined to distinguish their difference.Difference from the way reported previously to obtain antiserum with which ribulose-1,5-bisphosphate carboxylase(Rubisco)is purified from plant and then immune animal, eukaryotic expression vector of genetically engineered gene was used for preparation of antiserum against Rubisco in present study.Because Ribulose-1,5-bisphosphate carboxylase large subunit(rbcL)which is encoded by chloroplast genes has high homology with other close-related species and have no intron, so it can be directly amplified from Eremopyrum orientale genomic DNA based on published sequence of Eremopyrum rbcL. RbcL was constructed in eukaryotic expressed vector-pcDNA3 to prepare DNA vaccine, and was constructed in prokaryotic expressed vector-pGEX4T-1 to prepare purified protein for strengthening effect. ELISA and Western results showed that the potency and specificity of antiserum were high and suitable for immunolocalization.The above antiserum against rbcL was used to locate the Rubisco in leaves of four plants through immunogold localization. Inspection under ultramicroscope showed that the distribution of immunogold particles was markedly difference between Carbon three plants and ephemerals. In Brasscica and Wheat, stroma of mesophyll chloroplasts was densely labeled with immunogold particles for Rubisco. In Eremopyrum orientale and Lepidium apetalum, stroma of mesophyll chloroplasts was scarcely labeled with immunogold particles, indicating a low Rubisco content in mesophyll chloroplasts, but the stroma of chloroplasts of BSC was densely labeled with immunogold particles for Rubisco. These results showed that the distribution of key photosynthetic enzyme-Rubisco in two ephemeral plants is different from Carbon three plant but similar with Carbon four plant.