Yan-chao Yin a,?,Xiao-dong Zhang a,?,Zhi-qiang Gao a,Ting Hu a,Lin Yang a,Zhi-xin Zhang a,Wen-dong L ib*,Ying Liua,*.Over-expressing root-specific β-amyrin synthase gene increases glycyrrhizic acid content in hairy roots of Glycyrrhiza uralensis[J].Chinese Herbal Medicines (CHM),2019,11(2):192-199
根特异性过表达β-香树脂醇合成酶基因增加乌拉尔甘草毛状根中甘草酸的含量
Over-expressing root-specific β-amyrin synthase gene increases glycyrrhizic acid content in hairy roots of Glycyrrhiza uralensis
  
DOI:10.1016/j.chmed.2019.03.001
中文关键词:  β-香树脂醇合成酶  乌拉尔甘草  毛状根  根特异性  过表达
英文关键词:β-amyrin synthase  Glycyrrhiza uralensis Fisch.  hairy roots  root-specific  over-expression
基金项目:
Author NameAffiliation
Yan-chao Yin a,? aSchool of Life Sciences, Beijing University of Chinese Medicine, Beijing 102401, China 
Xiao-dong Zhang a,? aSchool of Life Sciences, Beijing University of Chinese Medicine, Beijing 102401, China 
Zhi-qiang Gao a aSchool of Life Sciences, Beijing University of Chinese Medicine, Beijing 102401, China 
Ting Hu a aSchool of Life Sciences, Beijing University of Chinese Medicine, Beijing 102401, China 
Lin Yang a aSchool of Life Sciences, Beijing University of Chinese Medicine, Beijing 102401, China 
Zhi-xin Zhang a aSchool of Life Sciences, Beijing University of Chinese Medicine, Beijing 102401, China 
Wen-dong L ib* bDepartment of Antibiotic, Beijing Institute for Drug Control, Beijing 102206, China 
Ying Liua,* aSchool of Life Sciences, Beijing University of Chinese Medicine, Beijing 102401, China 
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中文摘要:
      目的:乌拉尔甘草(Glycyrrhizia uralensis)是中国最常用的中药材之一,主要在中国种植。然而,栽培的乌拉尔甘草中甘草酸含量远低于中国药典规定的最低标准。在本文中,我们采用过表达β-香树脂醇合成酶基因(GuBAS)的方法来增加甘草酸含量。 方法:构建根特异性过表达GuBAS基因的植物表达载体,并以电转法转化到发根农杆菌中构建发根农杆菌工程菌。重组发根农杆菌工程菌侵染乌拉尔甘草的胚轴和子叶以诱导毛状根。诱导获得的毛状根经过培养后,采用HPLC法测定甘草酸含量。 结果:PCR和测序结果均显示获得了3个转基因毛状根系。通过qRT-PCR测定不同转基因毛状根系中的GuBAS拷贝数设为3,7和4。通过HPLC检测甘草酸含量,结果显示甘草酸仅存在于三个转基因毛状根系中,野生型毛状根中未测得。 结论:根特异性过表达β-香树脂醇合成酶基因增加了乌拉尔甘草毛状根中甘草酸的含量。
英文摘要:
      Objective: Glycyrrhizia uralensis, one of the most widely-used traditional Chinese medicines, is mainly cropped in China. However, many cultivars are less in glycyrrhizic acid than Chinese Pharmacopoeia requires. In this paper, we improved glycyrrhizic acid by regulating β-amyrin synthase gene (GuBAS). Methods: Tobacco root-specific promoter TobRB7 and GuBAS cDNA were obtained and combined with linearized pCAMBIA1305.1 to construct root-specific plant expression vector which was later transformed into Agrobacterium rhizogenes ACCC10060 by electrotransformation. The cotyledons and hypocotyls of G. uralensis were infected by the recombinant A. rhizogenes ACCC10060 to induce hairy roots. The GA content was quantified by HPLC. Results: The PCR and sequencing results both showed that three transgenic hairy root lines were obtained. The copy number of GuBAS in these transgenic hairy roots was intended by qRT-PCR to be 3, 7, and 4. GA was detected in the three transgenic hairy roots but in wild hairy roots. Conclusion: Over-expressing GuBAS root-specifically in hairy roots of G. uralensis enhanced GA accumulation.
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