水芹肉桂醇脱氢酶基因的克隆与表达特性分析

Cloning and Expression Characteristics Analysis of OjCAD Gene in Oenanthe javanica

以‘八卦洲水芹’及其紫色叶柄突变型水芹‘南选八卦洲紫水芹’为实验材料,利用RT-PCR方法从‘南选八卦洲紫水芹’中克隆得到水芹肉桂醇脱氢酶(CAD)基因,命名为OjCAD。OjCAD基因开放阅读框长为1 074bp,编码357个氨基酸。OjCAD蛋白相对分子质量为39 143.10,理论等电点为6.91,属于MDR家族。系统进化分析显示,水芹OjCAD与同属伞形科的胡萝卜CAD进化关系最近,具有高度保守性。OjCAD编码的蛋白属于疏水蛋白,空间结构主要由7个α-螺旋和17个β-折叠组成。实时定量PCR分析显示,OjCAD基因在紫色和非紫色水芹的叶片和叶柄中相对表达量存在差异,水芹OjCAD基因在叶片中的表达量显著高于叶柄,在‘八卦洲水芹’中的表达量高于‘南选八卦洲紫水芹’。该研究结果为进一步分析水芹木质素生物合成奠定了基础。

In this study, ‘Baguazhou Shuiqin＇and the purple-petiole mutant ‘Nanxuan Baguazhou Zishuiqin＇were used as plant materials.A gene encoding cinnamyl alcohol dehydrogenase（CAD）was cloned from the cDNA of‘Nanxuan Baguazhou Zishuiqin＇by RT-PCR method and designated as OjCAD.The OjCADgene contained a full-length open reading frame of 1 074 bp,which encoded 357 amino acids.The relative molecular mass and theoretical isoelectric point of OjCAD protein was 39 143.10 and 6.91,respectively.The protein encoded by OjCADis a member of MDR superfamily.The protein belonged to hydrophobic protein and its tertiary structure included 7α-helix and 17β-turn.Alignments and phylogeneticanalysis indicated that OjCAD was highly conserved and had the closest phylogenetic relationships with the CAD protein from Daucus carota.Quantitative real-time PCR analysis demonstrated that the relative expression levels of OjCAD gene in leaf blades and petioles of purple and non-purple Oenanthe javanica were different.The relative expression levels of OjCADgene in leaf blades were higher than that in petioles.The relative expression levels of OjCAD gene in ‘Baguazhou Shuiqin＇were higher than that in‘Nanxuan Baguazhou Zishuiqin＇.Those results provided a basis for further research on the lignin biosyn-thesis in O.javanica.