目的 在厚朴转录组测序基础上,对厚朴1-脱氧-D-木酮糖-5-磷酸合成酶(DXS)基因DXS1(MoDXS1)和厚朴DXS2(MoDXS2)基因进行全面的生物信息学分析。方法 利用生物信息学方法对厚朴MoDXS1和MoDXS2基因编码氨基酸序列的理化特性、亲/疏水性、功能域、二级结构、三级结构和系统发育进化等进行分析和预测;采用实时荧光定量PCR分析厚朴MoDXS1和MoDXS2基因的相对表达量。结果 MoDXS1和MoDXS2均为不稳定的亲水蛋白,蛋白结构域分析显示MoDXS1、MoDXS2与其他植物的DXS基因有很高的同源性;二级结构均为混合型结构的蛋白质,α-螺旋是所有基因多肽链中大量的结构元件,利用同源建模法对三级空间结构进行了预测;同源序列对比显示MoDXS家族与烟草、丹参和拟南芥的DXS蛋白具有较高的同源性;系统进化树分析表明MoDXS1与其他被子植物亲缘关系较近,但MoDXS2单独聚为一支;实时荧光定量PCR结果显示DXS1在厚朴与凹叶厚朴间无显著差异,但DXS2在凹叶厚朴中的表达量要明显高于厚朴;GC-MS检测结果表明,3种指标性成分β-石竹烯、β-氧化石竹烯和β-桉叶油醇在凹叶厚朴皮或叶中的量均高于厚朴。结论 本研究结果为解析厚朴萜类次生代谢调控机制奠定了理论基础。

Objective Based on the data of transcriptome sequencing of Magnolia officinalis, MoDXS1 and MoDXS2 genes were completed in detail by using bioinformatics methods. Methods MoDXS1 and MoDXS2 genes were analyzed and predicted by the tools of bioinformatics in the following aspects: physical and chemical characteristics of amino acid sequences, function domain, hyophobicity or hydrophilicity, secondary structure and tertiary structure of protein, molecular phylogenetic evolution, and so on; The expression levels of MoDXS1 and MoDXS2 were identified by real-time PCR. Results ORF Finder indicated that MoDXS1 and MoDXS2 genes were full-length, and they all were unstable hydrophobic proteins; Structural domain of MoDXS1 and MoDXS2 showed high homology with other plants; The secondary structures all were hybrid architecture, and alpha helixes were the major motifs, tertiary structure of protein was predicted by Homology modeling; Sequence alignment that MoDXS family had relative close relationship to the DXS of Nicotiana tabacum, Salvia miltiorrhiza, and Arabidopsis thaliana; The results of evolutionary relationship analysis showed that MoDXS1 had relative close relationship to angiosperm, but MoDXS2 was clustered in a clade solely; The expression levels of DXS1 in M. officinalis and M. officinalis var. biloba were not significantly different, but the expression level of DXS2 in M. officinalis var. biloba was higher than that in M. officinalis. By applying the technique of GC-MS, the contents of major volatile components β-caryophyllene, β-caryophyllene oxide, and β-eudesmol in M. officinalis var. biloba are higher than those in M. officinalis. Conclusion The results provide theoretical reference for studies on secondary metabolic regulation in terpenoid of M. officinalis.

国家“863”计划项目(SS2014AA022201)