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通过外源添加茉莉酸甲酯(MeJA),研究其调控液体培养条件下人参发状根中皂苷生物合成途径相关酶基因在诱导子作用时表达情况。以四年生吉林人参主根诱导的人参发状根无性系为材料,利用香草醛-硫酸比色法测定MeJA处理前后人参发状根的总皂苷含量;同时,利用荧光实时定量PCR测定MeJA处理前后人参发状根中鲨烯合成酶(squalene synthase,SQS)、鲨烯环氧酶(squalene epoxidase,SQE)、氧化鲨烯环化酶(oxidized squalene cyclase,OSC)、达玛烷二醇合成酶(dammarenediol synthase,DS)、β-香树脂合成酶(β-amyrin synthase,β-AS)和环阿屯醇合成酶(cycloartenol synthase,CAS)基因的相对表达量。结果表明:获得MeJA添加到人参发状根中最佳添加浓度为6×10~(-4)μmol·L~(-1)、添加时间为22 d、作用时间为5 d;MeJA的添加可以提高人参发状根中过氧化物酶(PPD)、过氧化氢酶(CAT)和过氧化物酶(PPD)的酶活性;经过MeJA处理后人参发状根中人参皂苷生物合成途径的SQS,SQE,OSC,DS,β-AS基因的表达变化均有显著提高,然而CAS基因的表达变化不显著。因此说明在添加MeJA处理后,皂苷生物合成途径中SQS,SQE,OSC,DS,β-AS基因表达情况与PPD,CAT,PPO的酶活性的变化趋势也相一致。
Exogenous addition of methyl jasmonate (MeJA) to study the liquid culture conditions under the regulation of ginseng hairy root saponin biosynthetic pathway related enzyme gene expression in the role of elicitors. Mesenteric rhizogenes induced by MeJA was used as raw material to measure the total saponin content of the roots of Panax ginseng and MeJA before and after treatment with MeJA. Meanwhile, the contents of ginseng root before and after treatment with MeJA were determined by real-time fluorescence quantitative PCR Hairy root squalene synthase (SQS), squalene epoxidase (SQE), oxidized squalene cyclase (OSC), dammarenediol synthase, DS, β-amyrin synthase (β-AS) and cycloartenol synthase (CAS) The results showed that the optimum concentration of MeJA added to the hairy roots of Ginseng was 6 × 10 -4 mol·L -1, the addition time was 22 days, and the action time was 5 days. The addition of MeJA Improve the enzyme activities of peroxidase (PPD), catalase (CAT) and peroxidase (PPD) in the roots of ginseng hairy roots; SQS of the ginseng saponin biosynthesis pathway in the hairy roots of ginseng after MeJA treatment, The expression of SQE, OSC, DS and β-AS gene were significantly increased, however, the expression of CAS gene was not significantly changed. Therefore, after addition of MeJA, the trend of SQS, SQE, OSC, DS, β-AS gene expression and the enzyme activities of PPD, CAT and PPO in the biosynthetic pathway of saponin were also consistent.