Studies On Establishment Of Tissue Culture And Cell Suspension Culture Systems Of Dendranthema Indicum Var. Aromaticum

Dendranthema indicum var. aromaticum is a kind of the Chrysanthemum in composite family, it is one new wild resources. Its flowers, leaves and roots have the rich of fragrance and there has tens of thousands of it in the golden autumn, very beautiful.sometimes folks often make sachets by the shade desiccated flowers and leaves. The plant contains some terpene compound, for example:Alpha-the thujone, Beta-the thujone and the Borneo camphor and so on, and it widely used in the industries for chemical, drink and medicine. At present Dendranthema indicum var. aromaticum is a new of medicinal plant resources be worth researching, developing and using, because its flowers and leaves have the effect of clearing heat and detoxicating. Now this thesis has mainly discussed the tissue culture of the wild plant Dendranthema indicum var. aromaticum and the establish of its cell suspension system,meanwhile afford the theory basis for the protoplast separation, purification and the protoplast fusion of it. So the mainly research results are as follows:1. The experiment of use different combinations of plant growth regulator 6-BA and NAA to induce the cluster buds of Dendranthema indicum var. aromaticum indicate that:its soft stem is easier induce, the highest inductivity may attain 90% with the culture medium includes 6-BA 0.1 mg/Land NAA 0.1 mg/L, and grow well, the average of each exophyte can engender 8.7 buds. The optimum culture medium of subculture cluster buds is MS+6-BA 0.3 mg/L+NAA0.1 mg/L. When disposed with different combination of 6-BA and NAA, all the Dendranthema indicum var. aromaticum can take root, but the best culture medium is MS+0.3mg/LNAA, the longest root can reaches 5.8cm, the most quantity reaches 10 and strongest. In the other medium, the root is longer, thiner and the rooting rate lower.2. The result of useing different floral organs as explant through the approach of callus to establish sterile cultivate system shows that:the best substrate of petals callus induction is MS+NAA 0.2 mg/L+6-BA 1.5 mg/L; and the best of torus callus induction is MS+6-BA 2.0 mg/L+NAA 0.2 mg/L. Afterwards tissue culture different developmental of Dendranthema indicum var. aromaticum torus as explant, successful induction callus and the better effect is the time early in budding show. In the same period of torus,the inductivity decreased with the increase of diameter, the diameter more smaller, the rate of callus more faster and the effect of induction more better. The best substrate of peduncle callus induction is MS+ NAA 0.1 mg/L+6-BA 1.0 mg/L+2,4-D 2.0 mg/L; and the best of anther is MS+6-BA 2.0 mg/L+2,4-D 1.0 mg/L with the concentration of cane sugar 6%, Later transfer it to the substrate of MS+6-BA 0.5 mg/L+NAA 0.2 mg/L and the most adventitious bud out. Last the callus induction of all kinds of flowers organs as explant rank in order from easy to hard is torus>anther>peduncle>petals. When use NAA may enhance the effect of rooting, the better concentration is 0.3 mg/L, transplanting the survival rate was80%.3.There is tremendous influence on the callus induction of different Dendranthema indicum var. aromaticum explants, the result show that the stems is best whether among the inductivity, the growing trend and the degree, although the inductivity is not highest, but the degree is lowest and growing better than the other. The inductivity achieves 100% when the substrate is MS+ BA 0.2 mg/L and 2,4-D 1.0 mg/L. The substrate is MS+NAA 0.1 mg/L+6-BA 0.5 mg/L, the callus induce bud be highest, reaches 86.7%.4.The best callus suspension culture substrate is MS+BA 0.2 mg/L+2,4-D 0.5 mg/L, the increment be biggest and cultivate 2 g callus after 18d, the fresh weight will achieve 8.12 g. In the cell suspension culture process, the fresh weight, PH and the sucrose density has had certain change, its growth curve assumes "S" curve, there existence the growth lag phase, the logarithm vegetal period, the plateau; and the pH value's change presents continuously the declining trend, but may maintain above 4.0, also may maintain the acidity for cell growth; The sucrose density presents the declining trend too, because the cell growth process need consume much sugar.