College of Food Engineering and Nutritional Science, Shaanxi Normal University, Xi'an, China.
J Food Biochem. 2019 Nov;43(11):e13016. doi: 10.1111/jfbc.13016. Epub 2019 Aug 26.
Sour rot, caused by Geotrichum citriaurantii (G. citri-aurantii.), is one of the most serious grapevine secondary infection diseases in China. We have determined that pterostilbene, the most important phytoalexin, effectively inhibits the activity of G. citri-aurantii. To study the synthesis mechanism of pterostilbene against G. citri-aurantii in grape, we initially detected the content of pterostilbene present in grapes infected by G. citri-aurantii with the use of UHPLC-QQQ-MS . Pterostilbene levels are controlled by the resveratrol O-methyltransferase (ROMT) gene, and within grape samples is positively related to the accumulation of resveratrol. The pPIC9k-ROMT vector and plasmid pCAMBIA2300-GFP-ROMT were constructed for expression purposes. The pPIC9k-ROMT vector was expressed in Pichia pastoris GS115 and plasmid pCAMBIA2300-GFP-ROMT was expressed in onion. Analysis of qRT-PCR amplification samples revealed that gene expression was induced rapidly in grape as a defense against G. citri-aurantii. Western blot analysis verified that the recombinant protein was successfully expressed. PRACTICAL APPLICATIONS: Pterostilbene is a trace and efficient phytoalexins produced by plant' secondary metabolism, which exhibits good pharmacological activity. As an important protective agent in plants, it can improve the antioxidant capacity and resistance to adversity of plants. However, the method which could be used for mass production of pterostilbene has not been reported currently. The key gene of pterostilbene biosynthesis was investigated and verified in this paper, which provides the theoretical basis for the industrial production of pterostilbene. The study of pterostilbene is significant for the prevention and treatment of G. citri-aurantii disease, and has important practical applications for the development and production of pesticides.
酸腐病由桔青霉（Geotrichum citriaurantii）引起，是中国葡萄园中最严重的二次感染病害之一。我们已经确定，植物抗毒素白藜芦醇能有效抑制桔青霉的活性。为了研究葡萄对白藜芦醇桔青霉合成机制，我们最初使用 UHPLC-QQQ-MS 检测感染桔青霉的葡萄中白藜芦醇的含量。白藜芦醇水平受白藜芦醇 O-甲基转移酶（ROMT）基因控制，与葡萄样本中白藜芦醇的积累呈正相关。为了表达目的，构建了 pPIC9k-ROMT 载体和质粒 pCAMBIA2300-GFP-ROMT。pPIC9k-ROMT 载体在毕赤酵母 GS115 中表达，质粒 pCAMBIA2300-GFP-ROMT 在洋葱中表达。qRT-PCR 扩增样品分析表明，葡萄中基因表达迅速诱导以抵御桔青霉。Western blot 分析验证了重组蛋白的成功表达。实际应用：白藜芦醇是植物次生代谢产生的微量、高效植物抗毒素，具有良好的药理活性。作为植物的重要保护剂，它可以提高植物的抗氧化能力和抗逆性。然而，目前还没有报道能够大规模生产白藜芦醇的方法。本文研究并验证了白藜芦醇生物合成的关键基因，为白藜芦醇的工业化生产提供了理论依据。研究白藜芦醇对防治桔青霉病具有重要意义，对农药的开发和生产具有重要的实际应用价值。
