Establishment And Optimization Of Tissue Culture System Fortaxus

Taxus is a rare medicinal gymnosperms, it contains Taxol which is a significant treatment effect for cancer. It is the best way to industrial production of paclitaxel to solve the paclitaxel drug source needs. By using the techniques of Taxus tissue culture, Taxus tissue cells were cultured in an artificial environment, screening of high taxol cell lines can achieve a large number of continuous production of the target product paclitaxel. It is the key of culture technology to produce paclitaxel that establish the taxus tissue culture system, through this stage of the Taxus tissues and organs training, having access to stable growth and proliferation of cells, established cell lines for the screening of high taxol. In this experiment, taxus tissue culture conditions in the various influencing factors being compared and screened, the process of organization culture of Taxus about explants selection, explant inoculation and disinfection, media types in the callus induction, pH, hormone levels, anti-browning conditions were optimization. On this basis, induced20stable growth and proliferation of Taxus cell lines, the Taxus tissue culture system were initially established, laid the foundation for future screening of high-yielding cell lines, The results showed that:1、Endophytes in Taxus are fungi and bacteria, some taxus organization materials subjected to endogenous bacteria contamination in the tissue culture process. This experiment studied the way of the Taxus inhibition of endophytic bacteria in the tissue culture. The results showed:The Taxus endophytes was fungi, the Taxus endophytes were significantly inhibited by antifungal antibiotics. The maximum inhibitory rate of the endophytes was by using500mg L-1fluconazole, and organizational survival rate for100%.2、Disinfectant categories and disinfection of time exist affects on callus induction, The results showed:the explants were disinfected by0.1%mercuric chloride in12min, The rate of contamination was the lowest, the rate of Callus induction was100%. Similarly, the explants were disinfected by0.1%mercuric chloride+300mg·L antibiotic in6min and20min, the rate of Callus induction was100%.3、Screening of callus induction, proliferation stage of medium type and composition, tissue cell culture environment. The results showed callus induction of the optimum medium was for:Improvement of B5+TDZ0.002+NAA1.0+2,4-D1.0+100mg·L-1ascorbic acid, pH=6,25℃, dark. The best medium of taxus organizations proliferation was for:Improvement of B5+TDZ0.002+NAA0.5+2,4-D0.5+100mg·L-1ascorbic acid, pH=6,25℃, dark.4、20genotype stability induced by growth and proliferation of Taxus cell lines were established. The results showed taxus cells color to yellow, light yellow, more fluffy, sand granular had subculture rate, fresh weight of Cell increased fast in unit time.