紫薇属香叶醇生物合成途径关键基因分离与功能分析

Breeding fragrant cultivars is one of the most important goals of Lagerstroemia indica (crape myrtle) breeding. L. caudata is the fragrant germplasms in the genus of Lagerstroemia. The lack of depth study of its fragrant traits resulted in the slow progress of aroma crape myrtle breeding. Our previous study found that the main different flower volatiles between L. caudata and crape myrtles were geraniol, neral and citral. Neral and citral are the oxidation products of geraniol. Therefore, geraniol synthesis and metabolism may be the key part, which led to the difference of flower odor. To explore the biological pathways of geraniol synthesis in Lagerstroemia, L. caudata and ‘Baiyun Yingxia’ were used as materials. On metabolic level, geraniol volatilization and temporal volatile in flowers were analyzed using semi-quantitative methods. Endogenous quantity of geraniol was determined by GC-MS. The catalytic efficiency of flowers was analyzed by adding GPP and geraniol to crape myrtle as exogenous substrates. On gene level, genes related to geraniol synthase (GES) were isolated based on the known sequences. Analysis of gene structure and phylogeny, expression of GES in Escherichia coli, enzyme assays, alternative substrate analysis, real-time PCR were studied gradually to found the key GES genes. Then, Arabidopsis thaliana and Nicotiana benthamiana were used as model plant for functional verification. Yeast one-hybrid method was also used to detect transcription factors (TFs) of GES. The results will finally explain the reason of the lack of aroma and elucidate the molecular mechanism of geraniol biosynthesis in Lagerstroemia, which have an important effect on geraniol biosynthesis and regulatory mechanisms of ornamentals and lay the foundation for aroma molecular breeding in Lagerstroemia.

培育香花紫薇品种一直是紫薇育种的重要目标。尾叶紫薇是香花种质资源，但由于缺少对其花香性状的深入研究，紫薇香花育种进展缓慢。课题组前期研究发现尾叶紫薇与紫薇主要挥发成分差异为香叶醇和其氧化产物橙花醛、柠檬醛。因此，香叶醇的合成与代谢可能是二者花香差异的关键环节。为探索紫薇属香叶醇生物合成途径，本研究以尾叶紫薇与紫薇品种‘白云映霞’为材料，一方面，研究花中香叶醇挥发量和时空挥发情况，比较花中香叶醇内源含量，分析紫薇对外源底物的催化效果；另一方面，克隆香叶醇合酶基因，进行生物信息学分析，开展原核表达与酶催化底物筛选，分析基因时空表达情况，筛选香叶醇生物合成关键基因，转化模式植物进行功能验证，通过酵母单杂交探索该基因的调控因子。最终揭示紫薇花香缺失的原因，明确紫薇属香叶醇生物合成的分子机理。研究结果将对阐明观赏植物香叶醇生物合成与调控机制有重要意义，为紫薇香花分子育种奠定基础。

为探索尾叶紫薇与紫薇花香差异分子机理，本项目通过对3组12种化合物的比较分析，筛选出适用于尾叶紫薇香气物质分离、浓缩的前处理方法，建立了适于紫薇花挥发物测定的外标半定量技术体系，确定了尾叶紫薇香气物质的组成特征，研究了尾叶紫薇香气物质的释放机理。通过转录组和代谢组的联合分析，推测尾叶紫薇香气物质的主要合成途径，筛选出与主要香气物质合成调控相关的基因，并分别在尾叶紫薇和紫薇‘白云映霞’中进行验证。研究结果有助于揭示紫薇花香代谢调控的机理，对定向培育芳香宜人的紫薇新品种具有重要的指导意义。