龙脑樟法呢基焦磷酸合酶基因克隆及表达模式分析  

Molecular Cloning and Expression of Farnesyl Pyrophosphate Synthetase in Cinnamomum camphora

作　　者：杨琳[1] 张杭颖[1] 杨帆 朱泽榕 李秋妹 张君诚[1] YANG Lin;ZHANG Hangying;YANG Fan;ZHU Zerong;LI Qiumei;ZHANG Juncheng(Fujian Provincial Medical Plant Exploitation and Utilization Engineering Research Center,Sanming University,Sanming,Fujian 365004,China;School of Food Science,Fujian Agriculture and Forestry University,Fuzhou,Fujian 350000,China)

机构地区：[1]三明学院药用植物开发利用福建省高校工程研究中心,福建三明365004 [2]福建农林大学食品科学学院,福建福州350000

出　　处：《热带作物学报》2022年第10期1998-2005,共8页Chinese Journal of Tropical Crops

基　　金：福建省科技厅面上项目(No.2020J01382);福建省教育厅中青年教师教育科研项目(No.JAT190703)。

摘　　要：法呢基焦磷酸合酶(farnesyl pyrophosphate synthetase,FPS)是萜类及其衍生物合成的关键限速酶。本研究以龙脑樟为材料,基于转录组测序,克隆CcFPS基因,利用生物信息学软件分析该基因编码的氨基酸序列特征,并采用实时荧光定量PCR(qPCR)检测CcFPS基因的表达,分析龙脑樟根、茎、叶中该基因的表达与总萜含量的关系。结果表明:CcFPS基因的开放阅读框为1053bp,编码350个氨基酸,分子量为40.4kDa,等电点pI为5.25,亲水性平均值(GRAVY)为-0.299。氨基酸同源性与沉水樟最高(98.86%),含有7个相同的保守结构域和2个富含天冬氨酸的基序[DDXX(XX)D]。二级结构α-螺旋、β-折叠、β-转角和无规则卷曲的比例分别为61.14%、6.57%、2.86%和29.43%,三级结构预测与沉水樟极其相似。系统发育树(NJ树)结果显示,CcFPS蛋白与同为樟科的沉水樟、山苍子的FPS处于同一个分支上。CcFPS基因在龙脑樟的根、茎、叶中均有表达,且表达具有组织特异性,在叶中的相对表达量最高,根中次之,茎中最低。在CcFPS蛋白调控的萜类物质代谢通路中,茎中的总萜含量最高,根中次之,叶中最低。龙脑樟FPS基因表达与总萜含量具有相关性,为进一步解析FPS基因在龙脑樟萜类化合物合成过程中的作用提供参考。Farnesyl pyrophosphate synthase(FPS) catalyzes the key step of isoprenoid biosynthesis. The expression level of FPS gene influences the composition and content of essential oil in Cirmamonun camphora. In this research,based on transcriptomic analysis, the open reading frame(ORF) sequence of CcFPS was cloned from C. camphora. The characteristics of amino acids of CcFPS protein was analyzed by bioinformatic softwares. The expression pattern of CcFPS and total terpene content were detected from leaf, root, and stem by quantitative real-time PCR and ultraviolet spectrophotometry, respectively. The results showed that the ORF of CcFPS was 1053 bp in length, encoding 350 amino acids with molecular weight 40.4 kDa, isoelectric point pI 5.05, and grand average of hydropathicity(GRAVY)-0.299.The secondary structure contained 61.14% α-helice, 6.57% extended strand, 2.86% β-turn and 29.43% random coil. The three-dimensional structural was predicted to be similar with FPS of C. micranthum. The CcFPS protein possessed seven conserved domains and two aspartate-rich motifs [DDXX(XX)D] and showed the highest homology with FPS of C.micranthum(98.86%). Phylogenetic tree analysis showed that the CcFPS protein was clustered into the same group with the FPS proteins from C. micranthum and Litsea cubeba. The expression of CcFPS showed tissue specific expression,highest level in leaf and lowest level in stem, although it’s detectable in leaf, root and stem. The total terpenes content was the highest in stem, following in root and the lowest in leaf. The total terpene content was correlated with the expression of CcFPS. The study would provide insights into understanding the role of CcFPS in the terpenoids metabolic in C. camphora.

关 键 词：龙脑樟 法呢基焦磷酸合酶(FPS) 基因表达 总萜 

分 类 号：Q949.7[生物学—植物学]