Induction And Liquid Culture Of Panaxnotoginseng Hairy Roots

Panax notoginseng(Burk.)F.H.Chen is a unique Chinese herbal medicine, a member of the genus Panax of the Araliaceae family. In this study, the hairy roots of Panax notoginseng were successfully achieved through infection of callus-derived explants by Agrobacterium rhizogenes. Since hairy roots have some merits of fast growth without addition of growth hormones, genetic stability and strong ability of secondary metabolites accumulation. It’ll be a promising approach to solve the problem of lack of Panax notoginseng material supply with the method of cellular engineering.First, the effects of antioxidants on the browning and growth of Panax notoginseng callus were examined; Secondly, establishment of culture system for regeneration of P. notoginseng plantlets via callus differentiation was carried out; last, attempts to induce P. notoginseng hairy roots were made by infection of explants of regenerated plantlets with various strains of Agrobacterium rhizogenes.The main results are as follows:1. Dry weight of cells was used as index, the formula of media for P. notoginseng callus growth was optimized through combination of 2,4-D and 6-BA. The results showed that the formula of MS+2,4-D 1.0 mg/L+ 6-BA 0.2 mg/L was suitable for P. notoginseng callus multiplication, and MS+2,4-D 2.0 mg/L+ 6-BA 0.5 mg/L for the induction of granular-like callus.2. Effects of antioxidants on the browning and growth of P. notoginseng callus showed that antioxidants caused inhibition on non-browning callus growth. Compared with that of citric acid, vitamin C led to better preventative and inhibitory effect on the browning of P. notoginseng callus; however, addition of citric acid could cause better accumulation of saponins than that of Vc.3. Effects of p H on the two states of callus showed that both of the best p H were 5.6.4. Effects of different culture period on culture callus showed that different culture period of callus slightly influent growth and browning of callus, so the best period was 21.5. The results of orthogonal experiment of different hormones concentration showed that the best culture medium for callus to induce adventitious buds was MS+ BR 1.0 mg/L+ KT 1.0 mg/L+ IBA 1.0 mg/L. The best sub-culture medium of adventitious buds was MS+ NAA 0.2 mg/L+ KT 0.5 mg/L. MS+ KT 1.0 mg/L+ IBA 0.2 mg/L was the best for adventitious roots inducted and regeneration plant from embryonic callus.6. Three different explants were infected by three different strains respectively, then hairy roots induced with different concentrations of AS when co-culture. The results showed that the best explant, Agrobacterium rhizogenes and concentration of AS were leaves of regeneration plant, strains of ACCC10060 and 20 mg/L.