| As a precious medicinal woody plant, Phellodendron amurense, is both important timber tree species and rare pharmacal plant. It’s rich in alkaloid, including Berberine, Jatrorrhizine, Palmatine and so on. Besides the long nature breeding cycle and the low breeding rate of woody plant, natural disaster and destroy by human all restrict wilding P. amurense’s using. Protect P. amurense resources more effectively and utilize them more rationally are becoming increasingly important. Rapid Propagation of P. amurense through tissue culture, which is not only shorten the period of growth but also retain good strains of seeds breeding. Improve the content of effective composition of P. amurense through tissue culture and cell culture technics has particular significance for protection of P. amurense resources and application in science fields. In this paper, an effective tissue culture regeneration system was established through using stems of sterile plantlet of P. amurense. Factors influencing shoot regeneration from stems including culture medium type, combinations of plant growth regulators and carbon source in the medium were investigated and physiological and biochemical indexes changes during regenerative plantlet rooting were discussed. Meanwhile, the effects of different color plastic films, carbon source and nitrogen on callus and on regenerative plantlet were systematically studied also. Following conclusions were got:1. Factors influencing shoot regeneration from stems were studied, the results showed MS medium supplemented with1.5mg/L6-BA and0.5mg/L NAA was the best for promoting shoot regeneration. The effects of different color plastic films on shoot regeneration from stem explants of P. amurense and on some physiological and biochemical indexes changes during shoot regeneration were investigated, and the result showed that blue films promoted shoot regeneration, improved antioxidant enzymes activity and soluble protein content. It was found that blue films induced higher level of chlorophyll content and were the best for growth of tissue and content of secondary metabolic product, and the worst were under green and red ones.2. We made a research into physiological and biochemical indexes changes during regenerative plantlet of rooting on basis of establishment of regeneration of P. amurense. It was shown that biomass and antioxidant enzymes activity was improved, soluble sugar and protein content was increased but starch decreased during rootage induction period. The content of Berberine and Palmatine had no obvious fluctuations, but Jatrorrhizine was increased. This trend of indexes changes in rootage induction period contrary to rooting stage, and three important alkaloid was increased quickly. 3. Using the callus of P. amurense which were induced by stems as explants to describe the effects of different carbon source and nitrogen on physiological and biochemical indexes changes. The results showed that20g/L sucrose improved callus growth and accelerated alkaloid synthesis.Compare to sucrose and fructose,20g/L glucose increased soluble sugar synthesis. Fructose was more suitable for accumulation of starch, but the difference between20,30,40g/L fructose effects was not clearly. Meanwhile, the research also indicated that increased the quantity of total nitrogen improved the accumulation of alkaloid and antioxidant enzymes activity but inhibited the growth of the culture.4. Using color plastic films and sucrose, we investigated the effects of different light qualities and sucrose concentration on growth and alkaloid synthesis of regenerative plantlet. The results were as follows:yellow films were more favorable to alkaloid accumulation and growth of regenerative plantlet. Blue films increased the activity of peroxidase(POD) and catalase (CAT), improved total chlorophyll. Green films promoted the activity of superoxide dismutase (SOD). However, soluble protein content had no obvious changes in different color plastic films. It was found that20g/L sucrose was more suitable for growth of regenerative plantlet. Three alkaloid were more than other sucrose concentration without sucrose, and there is no significant regular relation between sucrose concentration and soluble carbon and starch content... |
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