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茉莉酸甲酯诱导积雪草毛状根培养物中积雪草苷产量提高。

Enhanced production of asiaticoside from hairy root cultures of Centella asiatica (L.) Urban elicited by methyl jasmonate.

作者信息

Kim Ok-Tae, Bang Kyong-Hwan, Shin Yu-Su, Lee Min-Jeong, Jung Su-Jin, Hyun Dong-Yun, Kim Young-Chang, Seong Nak-Sul, Cha Seon-Woo, Hwang Baik

机构信息

Division of Ginseng and Medicinal Crop, National Institute of Crop Sciences, RDA, Suwon 441-857, South Korea.

出版信息

Plant Cell Rep. 2007 Nov;26(11):1941-9. doi: 10.1007/s00299-007-0400-1. Epub 2007 Jul 14.

Abstract

Transformed root ("hairy root") cultures have been shown to be a good model for the study of many secondary metabolites. However, economically important compounds such as asiaticoside and madecassoside are produced in insignificant amounts in the root of Centella asiatica (L.) Urban. To overcome this problem, C. asiatica was transformed using Agrobacterium rhizogenes strain R1000 that harbors pCAMBIA1302 encoding the hygromycin phosphotransferase (hpt) and green fluorescence protein (mgfp5) genes and the hairy culture was coupled with elicitation technique. Hairy roots were obtained at a frequency of up to 14.1% from a tissue junction between the leaf and petiole. Abundant hairy roots were observed when co-cultivation of the plant with A. rhizogenes was done for 7 days (36.1%). Transformation was confirmed by PCR and Southern blot analyses. Five weeks after inoculation, no asiaticoside was detected in the hairy root samples. However, when 0.1 mM methyl jasmonate (MJ) was applied as an elicitor to the culture medium for 3 weeks, a large quantity of asiaticoside was generated (7.12 mg/g, dry wt). In the case of gene expression, 12 h after MJ treatment the expression of the CabAS (C. asiatica putative beta-amyrin synthase) gene in the hairy roots is significantly different from that of the control and this level of transcripts was maintained for 14 days. Our results showed that production of C. asiatica hairy roots could be optimized and the resulting cultures could be elicited with MJ treatment for enhanced production of asiaticoside.

摘要

转化根(“毛状根”)培养物已被证明是研究许多次生代谢产物的良好模型。然而,积雪草中经济上重要的化合物如积雪草苷和羟基积雪草苷的产量微不足道。为克服这一问题,利用携带编码潮霉素磷酸转移酶(hpt)和绿色荧光蛋白(mgfp5)基因的pCAMBIA1302的发根农杆菌R1000菌株对积雪草进行转化,并将毛状根培养与诱导技术相结合。从叶和叶柄之间的组织连接处获得毛状根的频率高达14.1%。当植物与发根农杆菌共培养7天时,观察到大量毛状根(36.1%)。通过PCR和Southern印迹分析确认了转化。接种五周后,在毛状根样品中未检测到积雪草苷。然而,当向培养基中添加0.1 mM茉莉酸甲酯(MJ)作为诱导剂处理3周时,产生了大量积雪草苷(7.12 mg/g,干重)。在基因表达方面,MJ处理12小时后,毛状根中CabAS(积雪草假定的β-香树脂醇合酶)基因的表达与对照有显著差异,且该转录水平维持了14天。我们的结果表明,积雪草毛状根的产量可以优化,并且通过MJ处理可以诱导所得培养物以提高积雪草苷的产量。

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