Study On Tissue Culture Of Polygonum Multiflorum Thunb

For the past few years, medicinal resources of Polygonum multiflorum Thunb were in short supply because of the destruction of natural environment, unlimited exploiting and outdated culture technique. Therefore, how to made the wild species as cultispecies, increase their quality , improve breeding variety, and rapid propagation were become important research projects. In this paper, the propagation by tissue culture,the cultivation of plantlets,and the composition analysis were studied ,including the induction of explants, buds proliferation, callus induction and differentiation, root induction,transplantation of tube plantlets, field planting and harvesting tuber roots of Polygonum multiflorum Thunb.Using HPLC and spectrophotometer method, the contents of 2, 3, 5, 4′-stilbene glucoside, aloe-emodin and total flavones of Polygonum multiflorum tuber roots were determined.The tuber roots were harvested from four different years cultspecies, a wild species of yunnan, and five processing polygonum multiflorum thunb of different areas. These researches provided important references for tissue culture and rapid propagation, germplasm conservation, artificial cultivation of Polygonum multiflorum Thunb, which is of great value to meet the market demand. The test result indicates:1. MS medium was the optimum basic medium for of Polygonum multiflorum tissue culture.2. The buds was the best explant for the tissue culture of Polygonum multiflorum Thunb. 12min sterilization was suitable for the explants.3. As it has the similar function, white sugar could substitute for sucrose in tissue culture of Polygonum multiflorum Thunb.4. Buds proliferation may become the main pathway of tube plantlets propagation.5. The different combination of auxin and cytokinin had different impact on buds proliferation. The results showed that:in different concentrations of 6-BA and IAA, the proliferation coefficient of 6-BA1.5 mg/L +IAA0.8 mg/L is 7.6;in different concentrations of 6-BA and IBA, the proliferation coefficient of 6-BA1.5 mg/L +IBA0.3 mg/L is 8.8; in different concentrations of 6-BA and NAA, the proliferation coefficient of 6-BA1.0 mg/L +NAA0.025 mg/L is 6.8; in different concentrations of 6-BA,NAA and IBA, the proliferation coefficient of 6-BA0.75 mg/L +NAA0.01 mg/L+IBA0.1 mg/L is 6.4. Different kind of cytokinin had different effects on buds proliferation, 6-BA is more suitable than KT or ZT. Based on these data, it is found that the most suitable medium for bud proliferation was 6-BA1.5mg/L +IBA0.3 mg/L.6. The different basic medium had different impact on buds proliferation, the rank ordering was MS>B5>1/2MS>N6>ER; The proliferation coefficient could be improved by adding 5% coconut juice.7. Stem segment and buds were more suitable for callus induction of Polygonum multiflorum Thunb than leaves and root tuber . Callus induction under light is better than in dark. The best medium for callus induction was MS+6-BA0.5 mg/L +2,4-D1.5 mg/L. 5% juice extracted from the fruits of Siraitia grosvenorii was beneficial for callus induction; the impacts of different basic medium on callus induction was ranked as MS>ER>1/2MS>N6>B5.8. In the process of callus redifferentiation, buds could differentiated from different medium containing 6-BA3.0mg/L, 6-BA2.5mg/L+LFS0.1mg/L, and TDZ0.3 mg/L +NAA0.05 mg/L, among which TDZ0.3 mg/L +NAA0.05 mg/L was the best.9.The best culture medium for rooting was 1/2MS+IBA0.3mg/L+NAA0.05mg/L+0.2%AC. The best concentration of sugar for root induction was 2%.10. Of all the methods regarding acclimatization and transplants of seedling, without opening bottle cap or opening bottle cap three days were the best methods.11. The result shows that: The content of total flavones, 2, 3, 5, 4′-stilbene glucoside, aloe-emodin of wild tuber toots was higher than those of tissue cultured seedling. With the advancement of years, the total flavones, 2, 3, 5, 4′-stilbene glucoside, aloe-emodin of tissue cultured seedling in four different years increased accordingly. The contents of total flavones ,2, 3, 5, 4′-stilbene glucoside, aloe-emodin of processing polygonum multiflorum thunb in five different areas varied greatly. The contents from Henan and Hunan were relatively higher, while the contents from Guangxi,Sichuan and Guangzhou were relatively lower. The sequence of the effective composition's contents of Polygonum multiflorum Thunb cultivated in different years was three-year-old> two-year-old>one-year-old>half-year-old. The content of wild species of Yunnan was slightly higher than the content of Polygonum multiflorum Thunb cultivated for three years。Of these in five different areas,the quality of processing polygonum multiflorum thunb in Henan and Yunnan was better than that in Guangxi,Sichuan and Guangzhou.