Wax Synthesis And Stress Resistance Function Analysis Of Three WAX2Homologous Genes In Rice

The plant epicuticular wax plays an important role in adapting to external environment change and protecting plant against UV radiation, diseases and insect pests. Drought is becoming a severe problem in agricultural production. Studies have showed that the increase of epidermal wax content or some wax composition reduced non-stomatal water loss. In order to reveal the possible roles of Arabidopsis WAX2homolog genes in rice under different stresses, we have analysized the wax synthesis and stress resistance function of OsWAX2-1(LOC_Os06g44300), OsWAX2-2(LOC_Os02g08230) and OsWAX2-3(LOC_Os09g25850). The main results are as following:1. GFP protein fusion vectors of the three OsWAX2genes were constructed successfully and introduced into onion skin epidermal cells by Agrobacterium-mediate method. Through the sub-cellular localization, we found the fluorescence of OsWAX2-1-GFP fusion protein was observed in the nucleus and the fluorescence of OsWAX2-3-GFP fusion protein observed in the nucleus and plasma membrane.2. Drought treatment showed that overexpression of OsWAX2-1and OsWAX2-2in rice could increase survival rate of transgenic rice, but the survival rate of OsWAX2-3transgenic rice and OsWAX2-2-RNAi rice decreased compared to wild type plants. No significant differences between OsWAX2-1-RNAi and WT after drought stress were found.3. Investigation of high salt treatment showed that the shoot heights of Os WAX2transgenic seedlings were slightly different from WT. After200mmol/L NaCl stress,we found the root lengths of OsWAX2-1-OE and OsWAX2-2-OE were longer than WT and there were no significant difference between OsWAX2-3transgenic seedlings and WT.4. After stress treatment with ABA, we found OsWAX2-2transgenic seedlings are more sensitive than wild type and other transgenic plants.5. OsWAX2transgenic and wild type plants at flowering stage were subjected to drought stress by stopping watering. Physiological detection and biochemical analysis such as water loss rate, MDA content and chlorophyll leaching indicated that OsWAX2-l-OE and OsWAX2-2-OE displayed stronger tolerance than wild type plants. However, OsWAX2-3transgenic plants were sensitive to drought compared to wild type ones.6. Epicuticular wax crystalloids on leaves of different rice genotypes, including WT and transgenic rice, were observed by scanning electronic microscope. The result indicated that OsWAX2-1-OE and OsWAX2-2-OE had more epicuticular wax crystalloids than WT. And crystalloids distribution on the leaves of OsWAX2-1-RNAi and OsWAX2-2-RNAi were scattered. Compared to zhonghua, OsWAX2-2mutant showed decreased cuticular wax deposition. However, both OsWAX2-3-OE and OsWAX2-3-RNAi had less epicuticular wax crystalloids than WT.7. GC-MS analysis of the cuticular wax indicated that OsWAX2-1probably regulates biosynthesis of very long fatty acids, aldehydes and primary alcohol; OsWAX2-2probably regulates biosynthesis of C28and C30fatty acids; whereas OsWAX2-3probably plays an important role in regulating biosynthesis of very long fatty acids and primary alcohol.