AgNO3对颠茄毛状根生物碱及关键酶基因表达的影响

Influence of AgNO3 on tropane alkaloids and key enzyme genes in hairy roots of Atropa belladonna

关键词:  颠茄, 硝酸银, 托品烷类生物碱, 毛状根, 基因表达量

Key words:  Atropa belladonna, AgNO₃, tropane alkaloids, hairy roots, expression of gene

摘要:

为了研究不同浓度AgNO₃对颠茄毛状根生长过程中托品烷类生物碱及影响其合成关键酶基因表达的影响,该研究使用五种不同浓度的AgNO₃处理黑暗培养12 d的颠茄毛状根,2 d后收集毛状根并测定其鲜、干质量、托品烷类生物碱的含量、部分生理指标(MDA、Pro、可溶性糖、可溶性蛋白)、关键酶基因(pmt、trI、h6h)表达量。结果表明:AgNO₃虽然抑制了颠茄毛状根的生长,但却促进了托品烷类生物碱的积累,与对照相比,50、100、150 μmol·L^-1处理均显著性地提高了托品烷类生物碱的产量。同时在150 μmol·L^-1处理下,毛状根代谢途径中MDA与对照相比均具有显著性的提高,脯氨酸含量在150 μmol·L^-1下也有了显著性提高,达到了5.92 μg·g^-1FM,是对照的2.91倍。可溶性糖与可溶性蛋白含量在150 μmol·L^-1浓度下处理时其含量分别是对照的1.55倍和1.67倍。托品烷类生物碱合成过程中关键酶基因(pmt、trI、h6h)的表达量在不同浓度AgNO₃处理下均有不同程度的提高。由此可以推断,AgNO₃在150 μmol·L^-1处理下,可能通过调控脯氨酸、可溶性蛋白、可溶性糖这几种初级代谢产物或某几种关键酶基因的表达来影响颠茄毛状根托品烷类的合成。

Abstract:

In this study, we studied the influences of different concentrations of AgNO₃ on tropane alkaloids and key enzyme genes in metabolic pathway. We all know that the accumulation of plant secondary metabolites is directly related to some tprimary metabolism and activity of key enzymes. And AgNO₃ is a heavy metal, which can induce oxidative stress in plants and which has a great effect on plant growth and key enzymes of metabolism. Therefore, we added five concentrations of AgNO₃ into Atropa belladonna hairy roots, and cultivated in B5 liquid medium for 12 d, and then, we collec-ted hairy roots and determined fresh weight, dry weight, contents of tropane alkaloids, some physiological indexes included MDA, Pro, soluble sugar and soluble protein, genes expression level after cultivated for 2 d. The results showed that AgNO₃ promoted the accumulation of the alkaloids, although AgNO₃ inhibited the growth of hairy roots, 50, 100, 150 μmol·L^-1 AgNO₃ could improve the contents of tropane alkaloids compared with control. Meanwhile, 150 μmol·L^-1 treatment group significantly improved the hairy roots of the content of MDA compared with control. As well as the content of Pro increased to 5.92 μg·g^-1FM, 2.91 times of control. Then, the contents of soluble sugar and souble protein were significantly improved 1.55 and 1.67 times of control respectively by treating with 150 μmol·L^-1 AgNO₃ treatment. Thus, with the treatment of AgNO₃, gene expressions(pmt, trI, h6h) of key enzymes in the metabolic pathway of A. belladonna hairy roots all increased their produced effects. Therefore, we could be inferred that the contents of tropine alkaloids in A. belladonna hairy roots could regulate and control primary metabolism such as Pro, soluble protein, soluble sugar and secondary metabolism like expressions of key enzyme genes with the treating of AgNO₃. This study could provide the basic theories for study the synthesis mechanism of tropane alkaloids and an effective method to enhance the medicinal composition in the culture of hairy roots of Atropa belladonna in the future.