甘草毛状根诱导培养及其黄酮含量检测的研究

 目的 探讨影响甘草毛状根的诱导培养的因素及其总黄酮含量。方法 利用发根农杆菌的遗传和液体培养技术,研究了甘草（Glycyrrhiza uralensis毛状根的诱导和离体培养及其黄酮的产生情况。结果 不同发根农杆菌中,A4菌株侵染效果最好,约96%的子叶节外植体产生毛状根;不同外植体中,子叶节的转化效果最高,毛状根产生只需3~4 d;在毛状根的液体培养过程中,接种量为0.3 g,培养容积为500 mL时生长速率最快,毛状根湿重增长41倍;毛状根能产生药用成分甘草黄酮,根系中最高黄酮含量高于商品甘草,达干重的2.042%,约为未转化植株根的4.3倍;毛状根中的黄酮还分泌到培养液中,最高量为每100 mL培养液1.36 mg。结论 较为系统地研究了甘草毛状根的诱导培养条件和总黄酮量,为今后规模培养甘草毛状根生产药用甘草黄酮提供了可能性。

Study on Induction and in Vitro Cultivation of Glycyrrhiza uralensis Hairy Root and Its Flavonoids Production

OBJECTIVE To study the influencing factors of induction and cultivation of Glycyrrhiza uralensis hairy root and its flavonoids production. METHODS By using genetic transformation of Agrobactierium rhizogenes and liquid cultivation technique, the induction and cultivation of Glycyrrhiza uralensis hairy root and its flavonoids production were investigated. RESULTS Different inducement frequencies of hairy roots were found among different A. rhizogene strains. A4 exhibited the strongest infection ability with explants. The percentage of rooted cotyledonary node explants after infection was more than 96%. The inducement frequency from cotyledonary nodes was the highest among different explants. Hairy root could be initiated from cotyledonary node explants 3-4 d after inoculation with the strain of A. rhizogene A4. The largest growth rate during hairy root cultivation in vitro was shown with inoculum size of 0.3 g and volume of 500 mL. The wet weight of hairy root after liquid culture was 41 times higher than before. The hairy root could produce medicinal secondary metabolites, flavonoids, and the highest amount of flavonoids in the hairy root reached a level of 2.042% dry weight and was 4.3 times of those in the untransformed root. The highest amount of flavonoids in 100 mL liquid culture was 1.36 mg. CONCLUSION The cultivation conditions of Glycyrrhiza uralensis hairy root and the content of total flavonoids have been systematically established. The results presented here have made it possible for large scale cultivation of G. uralensis hairy root and production of flavonoids .