Study On Taxol Production In Taxus Chinensis Var. Mairei Cell Suspension Culture

Taxus chinensis var. mairei is a variation of Taxaceae Taxus in China, and it contains more Paclitaxel(Taxol) than the other plants of Taxus. But it is not nearly enough to meet market demand because of scarce wild resources of Taxus chinensis var. mairei and limited production capacity of Taxol. In recent years, studies on Taxus chinensis var. mairei tissue culture and cell culture have received extensive attention and the key points of those studies are selecting the explant, optimizing the medium, resolving the browning, establishing suspension system and optimizing the condition of production taxol. In this study, firstly, we selecte the explant; secondly, we optimize the medium; thirdly, we resolve the browning; lastly, optimize the condition of production taxol. The results are as follows:First, we selecte the suitable explant of inducing callus. We choose blade, stem without blade, stem with blade and mature embryo as explant, which were cultivated on the babylon medium with appropriate naphthylacetic acid and Benzyl Aminopurine. The results show that all of them could induce callus, but it is longer for blade and stem with blade to induce callus than mature embryo and stem without blade. The volume of the callus of the formers is smaller than the latters and the formers' inductivity is lower than the latters'. Considered all of the factors, we think the stem without blade is the best explant.Second, we optimize the medium of inducing callus. We choose mature embryo and stem without blades as explant, useing three kinds of medium and adding three kinds of hormone. The results show that the stems without blades grow better on babylon but the mature embryo grow better on1/2WPMSH. The better combination for the mature embryo is babylon added one milligram per litre naphthylacetic and half milligram per litre Benzyl Aminopurine. When we contract the structure of them, we found that the former's is compact and the latter's is loose.Third, the different additives'influence on callus browning. We choose mature embryo and stem without blades as explant, adding some active carbon, vitamin C or acid hydrolyzed casein. The results show that all of them could ease browning and the best one is active carbon and then is acid hydrolyzed casein. But when we add active carbon to the medium, the growth of the callus becomes lower than before which maybe because active carbon adsorbs the nutrient substance together with the polyphenol. At last, we consider acid hydrolyzed casein is better than active carbon.Last, we optimize the condition of production taxol. Use the callus induced by mature embryo and the stems without blades and with five times subculture to conduct cell suspension culture. The result shows that the initial concentration of cane sugar is thirty percent. During the whole process of cell suspension, the growth curve of the cells presents a model of "S"; when the cells are in stationary phase, the production of taxol in the suspension system is about thirteen microgramme per litre; we observe in the microscope and find that the status of the cell present rounded or elipse.