Abstract: To explore expression level of alkaloid and its key synthetase gene in Isatis indigotica upon Plasmodiophora brassicae exposure. The grades of disease severity according to morphology were verified. Moreover, histological observation, physiological and biochemical parameters have been collected together with transcriptomic and metabolomic analysis in Isatis indigotica after infection for 0, 7, 14, 21 d. The results were as follows:(1)After inoculation for 0, 7, 14, 21 d with Plasmodiophora brassicae, Isatis indigotica showed club root grades in 0, 1, 3, and 5 respectively, notably, cortical invasion occurred on 7 d.(2)When Plasmodiophora brassicae exposed lasting 14 d later, the contents of soluble protein and malondialdehyde, along with superoxide dismutase, peroxidase, polyphenol oxidase and catalase activities in Isatis indigotica increased significantly compared to control group at time depended manner.(3)A total of 161 alkaloids were detected in metabolomics, among those alkaloids, indoles were noticed as the most abundant form. There were 16, 17 and 39 discriminating metabolites had been spotted after infected with Plasmodiophora brassicae for 7, 14, 21 d, the most discriminating metabolites enriched at alkaloid and amino acid metabolism pathways.(4)Transcriptome analysis showed that there were 2 439, 256 and 6 437 genes expression alteration for 7, 14, 21 d compared to control, those differentially expressed genes enriched at 11 alkaloids related metabolism pathways. Markedly, expression level of 9 genes(encoding for enzymes thebaine synthase, tyrosine aminotransferase, indole-3-pyruvate monooxygenase and aldehyde dehydrogenase)were increased after infection for 7, 14, 21 d. The results reveal the interaction between Plasmodiophora brassicae and Isatis indigotica,explored the effects of Plasmodiophora brassicae on indole alkaloid synthesis and its key enzyme genes, and lay a foundation for later research on resistance genes and alkaloid secondary metabolic pathways in Isatis indigotica.

Key words: Isatidis Radix, Plasmodiophora brassicae, antioxidant enzymes, functional gene, indole alkaloid, metabolome

摘要： 为探究根肿菌胁迫对菘蓝生物碱及其合成关键酶基因表达的影响,该研究对根肿菌侵染后0、7、14、21 d的菘蓝进行病情形态分级、组织学观察、生理生化指标测定以及转录组学和代谢组学分析。结果表明:(1)接菌后0、7、14、21 d菘蓝根部分别发展为0级、1级、3级、5级的肿根,并且7 d是皮层入侵的关键时间。(2)接种根肿菌14 d后,菘蓝叶内可溶性蛋白、丙二醛的含量,超氧化物歧化酶、过氧化物酶、多酚氧化酶、过氧化氢酶的活性与同时间对照组比较显著提高,并随着接菌时间的延长呈增加的趋势。(3)代谢组学一共检测到161种生物碱,其中吲哚类生物碱数量较多; 与未接菌相比,菘蓝接菌后7、14、21 d分别存在16种、17种、39种差异代谢物且各组差异代谢物多富集在生物碱和氨基酸代谢通路。(4)转录组测序结果显示,与未接菌相比,菘蓝接菌后7、14、21 d分别存在2 439个、256个、6 437个差异表达基因,这3组共同富集到11个生物碱相关的代谢通路; 与未接菌相比,接菌后7、14、21 d有9个基因(编码4种酶THS、TAT、YUCCA、ALDH)表达量均上升。该研究结果揭示了芸薹根肿菌与菘蓝之间的互作机,探究了芸薹根肿菌对吲哚生物碱合成及其关键酶基因的影响,为后期菘蓝根肿病抗性基因及生物碱次生代谢途径的研究奠定了基础。

关键词: 板蓝根, 根肿菌, 抗氧化酶, 功能基因, 吲哚生物碱, 代谢组