杜仲MVA和MEP途径相关基因的亚细胞定位与表达分析

通过对杜仲基因组分析，筛选并克隆出MVA途径和MEP途径的相关基因全长（EuDXR，EuMCT，EuCMK，EuMDS，EuACOT，EuHMGS和EuHMGR），并通过生物信息学方法分析其结构特征，结果表明上述基因与其他已知物种相应基因的相似度达73%~85%。通过构建亚细胞定位表达载体，并瞬时转化烟草下表皮细胞后激光共聚焦显微镜下观察显示，EuDXR，EuMCT，EuCMK，EuMDS基因编码蛋白定位于叶绿体，EuACOT和EuHMGR基因编码蛋白定位于内质网，EuHMGS基因编码蛋白定位于细胞质膜。利用转录组测序技术分析上述基因的时空表达特性表明，MEP途径相关基因在杜仲叶片中大量表达，而MVA途径相关基因在杜仲幼果中大量表达，且杜仲幼果比叶片中的橡胶含量高，因此，推断MVA途径在杜仲橡胶合成中占主导作用。

Subcellular Localization and Expression Analysis of Genes from Eucommia ulmoides Involved in MVA and MEP Pathway

The full-long of EuDXR, EuMCT, EuCMK, EuMDS, EuACOT, EuHMGS and EuHMGR genes were isolated from Eucommia ulmoides. The structure characteristics were analyzed by using bioinformatics, which revealed that these deduced proteins had 73%-85% identities with the corresponding protein from other plants. Recombinant plasmid was introduced into the lower epidermal leaf explants of tobacco, and the transient expression of these genes with GFP fusion protein were observed with a Zeiss LSM. The subcellular localization analysis showed that the EuDXR, EuMCT, EuCMK, EuMDS protein were all located in the chlorophyll, EuACOT and EuHMGR protein were both located in the endoplasmic reticulum, EuHMGS protein was located in the plasma membrane. By RNA sequencing, the spatial-temporal expression patterns of these genes were detected. The expression of genes in MEP pathway is higher in leaves, but the expression of genes in MVA pathway is higher in young fruit. The rubber content in young fruit was higher than that in leaves. Therefore, the MVA pathway played a dominant role in the biosynthesis of Eu-rubber.