催化刺五加皂苷生物合成的法尼基焦磷酸合酶、鲨烯合酶和鲨烯环氧酶基因具有单核苷酸多态性和基因家族现象，但单纯的DNA序列变异无法全面揭示刺五加皂苷含量差异的分子机制。生物体中广泛存在DNA甲基化现象，其中启动子和编码区的甲基化对基因的表达调控等具有重要的作用。本项目拟在克隆和分析3个关键酶基因启动子和编码区的基础上，通过重亚硫酸盐测序法找到其中存在的DNA甲基化位点。进而利用不改变刺五加基因型，但可改变宿主DNA甲基化程度的内生青霉P116-1a诱导相同基因型刺五加形成不同程度的DNA甲基化。从而避免有性繁殖刺五加遗传背景复杂、无性繁殖刺五加DNA甲基化相同的弊端。结合各种甲基化状态下关键酶基因表达量及皂苷含量的测定结果，筛选出与皂苷含量密切相关的DNA甲基化位点，并将关键甲基化位点在野生刺五加中进行验证，从而基于关键酶基因启动子和编码区DNA甲基化解析刺五加皂苷含量差异形成的分子机制。

Farnesyl diphosphate synthase (FPS), squalene synthase (SS) and squalene epoxidase (SE) genes which catalyzed the biosynthesis of Eleutherococcus senticosus saponins all have single nucleotide polymorphism (SNPs) and gene family, but the difference of E. senticosus saponins content can not be fully revealed merely by the variation of the pure DNA sequence. DNA methylation is a common phenomenon in most organisms, among which the methylation of promoter and coding region plays an important role in the regulation of gene expression. This study is planned to find out the DNA methylation sites of promoters and coding regions of three cloned and analyzed key enzyme genes through bisulfite sequencing PCR (BSP). Penicillium minioluteum P116-1a is used to induce E. senticosus clones with various states of DNA methylation without changing the genotype of E. senticosus, thus the disadvantages of sexual reproduction that possibly lead to a complex genetic background can be avoided and the asexual reproduction E. senticosus had the same DNA methylation. Based on the results of saponins content and expression content of key enzyme genes under different states of methylation, we can screen out the specific methylation status that closely is related to saponins content. According to the confirmed key DNA methylation sites in wild E. senticosus, we can theoretically explain the molecular mechanisms of how DNA methylations of the key enzyme gene’s promoter and coding region lead to the variation of saponins content in E. senticosus.