Resveratrol, Plant Secondary Metabolite, Producted By Trangenetic Grape Cell With Agrobacterium Tumefaciens

Master degree dissertation (College of Life Science, Northwest University, China)Guo Bin (botany) Directed by Wei Ya-HuiResveratrol, plant secondary metabolite, producted by trangenetic grapecell with Agrobacterium tumefaciensResveratrol, stillbene compound,found in many plants,expecially in grape, shows to be active in artioxidant, anti-inflammatory and antioncogenic etc., But,the content of resveratrol is very little in various plants,which is not enough for people' requirment. So, the target of this investigation is to improve the productivity of resveratrol.In the course of investigationthree steps are included as follows. The first step is genetic transformation to grape cell with Agrobacterium.This step is to improve the grape cell's biomass increament .The second one is radiating by He-Ne Laser to the cell abtained through the first step. The key of this step ies to abtain a good strain which has a higher capability of producing resveratrol. And the last step is to induce the expression of RS in short time and prompt the composition of resveratrol by elicitors. Then the productivity of resveratrol increases dramaticly. At the same time, the optimal time of the adding of elicitors is screened.Three experiments are carried out with the callus of grape, and the experiment of genetic transformation with Agrobacterium tumefaciens 702 strain is the basis of the others. Trasgenic cell is obtained called MQ. The charicteristics of MO are that the production of cell is enhanced remarkably, though the production of resveratiol was equal to the control. The effects of several factors on transformation frequency is studied, and the result shows that transformation frequency is raised dramaticly by preculturing the callus with 0.4mol/L mannitol for 6h during preculture time (3day) before being infected with the 702 strain. And the Immaterial is taken as basal selection medium. In the condition, the transformation frequency is up to over 80% through paple electrophoresis.Then, the MO is radiated by He-Ne laser .A variety cell (N. T12) of secondary matter is bred by the first and second screen and studied on stability of mutantstrains. The yield of secondary matter (20-30)% than contrast. The shape and properties ofheredity of Ti2 is steady, at the same time, the period of production is sharply shortened.Besides, the effect of elicitors on the production of resveratrol is investigated in the last experiment. The results shows that the addition of AlCLs (7.5%)and botnis cinera (lOmg/L) into the material cell cultures of T^ at the final stage of fast-growing phase (the 30th day of culture) does not inhibited the cell growth, but dramatically increased resveratrol yield, which is as 4-5 times as control.