透明颤菌血红蛋白对黄芪甲苷生物合成的调控作用
Regulation of Vitreoscilla hemoglobin on biosynthesis of astragaloside IV
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摘要:
为了研究透明颤菌血红蛋白 (Vitreoscilla hemoglobin, VHb) 对黄芪甲苷生物合成的调控作用, 本文运用农杆菌介导法, 将透明颤菌血红蛋白基因 (Vitreoscilla hemoglobin gene, vgb) 导入黄芪毛状根中, 经PCR特征性引物扩增得到54个转基因株系。Southern blot分析, 证明vgb基因已经整合到转基因黄芪毛状根基因组中。RT-PCR分析结果表明, vgb基因在转录水平获得表达。转基因黄芪毛状根培养15天, 其生长量和增长倍数皆比非转基因黄芪毛状根高; 黄芪甲苷含量测定结果显示, 转基因黄芪毛状根黄芪甲苷含量是非转基因黄芪毛状根的5～6倍, 是山西产黄芪药材的10～12倍。研究结果表明, vgb基因的表达促进了转基因毛状根的生长, 提高 了黄芪甲苷含量。
Abstract:In the present study, the regulation of Vitreoscilla hemoglobin (VHb) on astragaloside Ⅳ biosynthesis was investigated. An intermediate expression vector consisting of the CaMV35S promoter fused to the vgb and nopaline synthase terminator was transferred into Astragalus membranaceus via Agrobacterium rhizogenes. The transgenic hairy roots were confirmed by PCR amplification and Southern blot hybridization. The expression of vgb in transgenic hairy roots was confirmed by RT-PCR. After 15 days cultivation, the dry weight and growth rate of transgenic hairy roots were higher than that of the non-transgenic hairy root. ELSD-HPLC analysis showed that astragaloside Ⅳcontent of transgenic hairy roots was 5 to 6 times of non-transgenic hairy root control and 10 to12 times of Radix Astragali from Shanxi Province. These results suggested that the expression of vgb promoted the growth of transgenic hairy roots, and increased the content of astragaloside Ⅳ.
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