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首页> 外文期刊>Acta Physiologiae Plantarum >Genetic transformation of Harpagophytum procumbens by Agrobacterium rhizogenes: iridoid and phenylethanoid glycoside accumulation in hairy root cultures
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Genetic transformation of Harpagophytum procumbens by Agrobacterium rhizogenes: iridoid and phenylethanoid glycoside accumulation in hairy root cultures

机译：发根农杆菌对原夏草的遗传转化：毛状根培养物中的烯类化合物和苯乙酮类糖苷积累

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A genetic transformation method using Agrobacterium rhizogenes was developed for Harpagophytum procumbens. The influence of three factors on hairy root formation was tested: bacterial strains (A4 and ATCC 15834), various types of explants and acetosyringone (AS) (200 μM). The highest frequency of transformation (over 50% of explants forming roots at the infected sites after 6 weeks of culture on Lloyd and McCown (WP) medium) was achieved using a combination of nodal stem explants and A. rhizogenes strain A4. The addition of 200 μM AS to root induction medium was found to enhance hairy root induction but its effect varied depending on bacterial strain and explant type. Three of the most vigorously growing hairy root clones of H. procumbens were chosen and analyzed for accumulation of iridoid and phenylethanoid glycosides. The transgenic nature of these root clones was confirmed by PCR amplification; they were positive for rolB and rolC genes. Harpagoside, verbascoside and isoverbascoside were identified by HPLC and LC–ESI-MS as the major compounds from all analyzed hairy root clones. The Hp-3 root clone showed the higher harpagoside content (0.32 mg g−1 dry wt.) compared with other analyzed transformed and non-tuberized untransformed roots of H. procumbens. However, the level of the compound in the hairy root clone was lower than that detected in a sample of commercially available root tubers of H. procumbens. The Hp-3 root clone also produced high amounts of verbascoside and isoverbascoside (8.12 mg g−1 dry wt. and 9.97 mg g−1 dry wt., respectively) comparable to those found in root tubers.
机译：开发了一种利用发根农杆菌的遗传转化方法来制备原夏草。测试了三个因素对毛状根形成的影响：细菌菌株（A4和ATCC 15834），各种外植体和乙酰丁香酮（AS）（200μM）。使用节点茎外植体和发根农杆菌菌株A4的组合，可以实现最高的转化频率（超过50％的外植体在Lloyd和McCown（WP）培养基上培养6周后在感染部位形成根）。发现在根诱导培养基中添加200μMAS可以增强毛状根的诱导，但其作用因细菌菌株和外植体类型而异。选择了三个最旺盛的毛鳞茎毛状根克隆，并分析了其类环烯醚酮和苯乙酮苷的积累。通过PCR扩增证实了这些根克隆的转基因性质。它们对rolB和rolC基因呈阳性。通过HPLC和LC-ESI-MS鉴定出了Harpagoside，马齿bas甙和异过山side甙是所有分析的毛状根克隆中的主要化合物。 Hp-3根克隆显示，与其他分析的原腐质酸根和未块茎化未转化根相比，Hp-3根的人参总苷含量更高（0.32 mg g -1 干重量）。但是，该化合物在毛状根克隆中的含量低于在商业上可获得的腐烂根瘤菌根茎样品中检测到的含量。 Hp-3根克隆也产生了大量的马齿side甙和异佛手甙（分别为8.12 mg g -1 干重和9.97 mg g -1 干重）。根块茎中发现的那些。

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