粘毛黄芩毛状根培养体系的建立及其黄芩苷的动态合成

Induction and culture of hairy roots in Scutellaria viscidula and its baicalin production

目的:建立粘毛黄芩毛状根培养体系,并对其生长特性和次生代谢产物的合成进行初步探索。方法:采用卸甲型根癌农杆菌Agrobacterium tumofaciens C58C1感染粘毛黄芩无菌苗的茎段,获得毛状根,并得到了优质株系;测定了毛状根的生长曲线;利用PCR对毛状根进行T-DNA转化的检测及利用HPLC进行黄芩苷含量检测。结果:PCR检测结果表明,发根农杆菌Ri质粒的rolC基因已整合入粘毛黄芩毛状根基因组中。C58C1感染粘毛黄芩无菌苗茎段8 d后,毛状根陆续在其伤口处产生,28 d时幼茎产生毛状根的外植体达81%。1/2 MS液体培养32 d后毛状根干重增加了17.42倍,总黄酮和黄芩苷含量分别增加了21.60,25.56倍。结论:粘毛黄芩毛状根离体培养的建立为进一步进行药用活性成分的工业化生产奠定了基础。

Objective： To estabhsh a culturing system of hairy roots of Scutellaria viscidula, and study the hairy roots growth and biosynthesis of flavonoid in the culturing system. Method： Hairy roots of S. viscidula were obtained from infected stem explants after infection with the disarmed Agrobacterium tumefaciens strain C58C1, elite strains were screened and growth curves were deter-mined. The transformation of Ri T-DNA was examined through PCR and baicalin content was examined through HPLC.. Result： Hairy roots appeared in vitro 8 days after inoculation of S. viscidula with disarmed A. tumefaciens strain C58C1. After 24 days the frequency of stems explants was up to 81%. Transformation was confirmed by the amplification of rolC genes from the hairy roots of S. viscidula. The results demonstrated that rolC genes could be expressed in hairy roots of S. viscidula. Under the 36 days suspension culture of S. viscidula hairy roots in 1/2MS medium, dry weight of hairy root increased 17.42 times, the content and baicalin increased 21.60 and 25.56 times. Conclusion： The establishment of the culturing system of hairy root of S. viscidula provided a foundation for further in- dustrial production of active drug component.