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作 者:吴天华 周骏辉[2] 赵玉洋 魏玉杰 陈芳 龚永福 袁媛[2] 黄璐琦 WU Tian-hua;ZHOU Jun-hui;ZHAO Yu-yang;WEI Yu-jie;CHEN Fang;GONG Yong-fu;YUAN Yuan;HUANG Lu-qi(h Academician Workstation,Jiangxi University of Traditional Chinese Medicine,Nanchang 330004,China;National Resource Center for Chinese Materia Medica,China Academy of Chinese Medical Sciences,Beijing 100700,Chinu;Gansu Academy of Agri-Engineering Technology,,Wuwei 733000,China)
机构地区:[1]江西中医药大学院士工作站,江西南昌330004 [2]中国中医科学院中药资源中心,北京100700 [3]甘肃省农业工程技术研究院,甘肃武威733000
出 处:《中国中药杂志》2021年第16期4111-4116,共6页China Journal of Chinese Materia Medica
基 金:国家科技基础性研究项目(2018FY100800);国家自然科学基金重大项目(81891013);中央本级重大支减项目(2060302);中央级公益性科研院所基本科研业务费专项(ZZ10-008)。
摘 要:血根碱是罂粟的主要活性成分,普罗托品-6-羟化酶(P6H)是血根碱生物合成途径上的基因之一,可把普罗托品氧化成6-羟基普罗托品。研究罂粟属药用植物P6H的基因多样性有助于获得酶活力高的P6H,提高血根碱生物合成的效率。该文利用罂粟属药用植物重测序数据,发现罂粟、鬼罂粟和虞美人P6H存在5种基因成员,并对其进行生物信息学分析;筛选伸长因子1α作为在根、茎中表达相对稳定的内参基因;通过实时荧光定量PCR检测了罂粟P6H基因在根、茎中的转录水平。从重测序结果中得到罂粟和鬼罂粟P6H各2种基因型,以及虞美人P6H的1种基因型。生物信息学分析显示3种罂粟属药用植物P6H蛋白均含P450超基因家族的保守结构域cl12078,均存在跨膜转运区,但是否存在信号肽有待验证;实时荧光定量PCR结果显示,罂粟P6H在根中的转录水平约为茎中的1.44倍,但两部位的P6H转录水平之间并无显著性差异。上述结果显示3种罂粟属药用植物普罗托品-6-羟化酶基因具有一定多样性,为罂粟中血根碱生物合成途径和机制研究提供依据。Sanguinarine is the main active component of the Papaver plants, and protopine-6-hydroxylase(P6 H), involved in the sanguinarine biosynthetic pathway, can oxidize protopine to 6-hydroxyprotopine. The investigation on the diversity of P6 H genes in the medicinal Papaver plants contributes to the acquirement of P6 H with high activity to increase the biosynthesis of sanguinarine. Five P6 H genes in P. somniferum, P. orientale, and P. rhoeas were discovered based on the re-sequencing data of the Papaver species, followed by bioinformatics analysis. With the elongation factor 1α(EF-1α), which exhibits stable expression in the root and stem, as the internal reference gene, the transcription levels of P6H genes in roots and stems of the Papaver plants were detected by real-time fluorescent quantitative PCR. As indicated by the re-sequencing results, there were two genotypes of P6H in P. somniferum and P. orientale, respectively, and only one in P. rhoeas. The bioinformatics analysis showed that the P6 H proteins of the three Papaver plants contained the conserved domain cl12078, which is the characteristic of p450 supergene family, and transmembrane regions. The existence of signal peptide remained verification. Real-time fluorescent quantitative PCR results revealed that the transcription level of P6 H in roots of P. somniferum was about 1.44 times of that in stems(α=0.05). The present study confirmed genetic diversity of P6 H in the three medicinal Papaver plants, which lays a basis for the research on the biosynthesis pathway and mechanism of sanguinarine in Papaver species.
关 键 词:罂粟 鬼罂粟 虞美人 重测序 普罗托品-6-羟化酶
分 类 号:S567.2[农业科学—中草药栽培]
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