甜叶菊毛状根培养系统的建立及绿原酸类成分的诱导

Establishment of Stevia rebaudiana Bertoni Hairy Root Culture System and Induction of Chlorogenic Acids

本文利用发根农杆菌A4诱导甜叶菊毛状根再生,建立毛状根生产绿原酸类物质的培养体系,并研究茉莉酸甲酯和水杨酸对毛状根中绿原酸类物质积累的影响。结果表明,经发根农杆菌A4侵染的甜叶菊叶片外植体在共培养14 d后诱导出了毛状根;PCR检测结果表明,发根农杆菌Ri质粒中rol B和rol C基因均已整合到毛状根中;毛状根在MS液体培养基中,培养到第14 d时分别加入不同浓度(50、100、200μmol/L)的水杨酸和茉莉酸甲酯进行诱导,处理第1、3、6 d后均抑制了毛状根的生长,但茉莉酸甲酯促进了毛状根中绿原酸类物质的积累,而水杨酸却抑制了毛状根中绿原酸类物质的积累。由此说明,发根农杆菌A4侵染甜叶菊可诱导出毛状根,该毛状根可用于绿原酸类物质的生产,茉莉酸甲酯可提高绿原酸类物质的含量。

Stevia rebaudiana hairy roots were induced using Agrobacterium rhizogenes A4. A culture system to produce chlorogenic acids in hairy roots was then established, and the effects of methyl jasmonate and salicylic acid on the accumulation of chlorogenic acids in hairy roots were studied. The results showed that Stevia rebaudiana hairy roots were successfully reproduced after 14 days＇ culture of Stevia rebaudianaleaf explants infected with Agrobacterium rhizogenes A4. Polymerase chain reaction（PCR） results indicated that genes rol B and rol C from Agrobacterium rhizogenes were integrated into the genome of Stevia rebaudiana. Hairy roots were cultured in Murashige and Skoog（MS） liquid medium for 14 d, and various concentrations（50, 100, and 200 μmol/L） of methyl jasmonate and salicylic acid were added to the cultures, respectively. After 1 d, 3d, and 6 d treatments, both plant elicitors inhibited the growth of hairy roots. However, methyl jasmonate promoted the accumulation of chlorogenic acids in hairy roots, and salicylic acid inhibited the accumulation of chlorogenic acids. Thus, Stevia rebaudianahairy root was induced by Agrobacterium rhizogenes A4 and could be used to produce chlorogenic acids; this could be promoted by methyl jasmonate elicitor.