内生真菌对刺五加皂苷合成关键酶基因表达及 皂苷含量的影响

目的:分析刺五加内生真菌菌株P116-1a,P116-1b,P109-4,P312-1对刺五加中皂苷合成关键酶基因SS(squalenesynthase),SE(squalene epoxidase),bAS(β-amyrin synthase)表达量和皂苷含量的影响。方法:伤口法回接内生真菌。以GAPDH为内参照基因,Real time PCR法检测关键酶基因表达量的变化。分光光度法测定刺五加的总皂苷含量。结果:回接P116-1a和P116-1b 30 d,刺五加SS基因的表达量显著提高(P<0.05),P109-4和P312-1无显著变化,之后出现降低-相等-降低的变化趋势。回接菌株P312-1 30 d,刺五加SE的表达量显著提升(P<0.05),回接菌株P116-1b和P312-1 90 d,SE的表达量分别显著提升至对照的130%,161%(P<0.05)。回接120 d,4个菌株处理的SE表达量均显著高于对照(P<0.05)。各处理bAS的表达量在回接60～120 d时均显著高于对照(P<0.05)。4个菌株均显著提高了刺五加的皂苷含量(P<0.05)。结论:菌株P116-1a,P116-1b,P109-4,P312-1可显著影响刺五加SS,SE,bAS的表达量,进而影响刺五加的皂苷含量,其中bAS为主要靶点基因。

Effect of endophytic fungi on expression amount of key enzyme genes in saponins biosynthesis and Eleutherococcus senticosus saponins content

Objective: To analyze the effect of endophytic fungi on expression amount of key enzyme genes SS(squalene synthase gene),SE(squalene epoxidase gene) and bAS(β-amyrin synthase gene) in saponin biosynthesis and saponins content in Eleutherococcus senticosus.Method: Wound method was used for back meeting the endophytic fungi to E.senticosus.With GAPDH as internal control gene,the expression of key enzyme genes was detected by real time PCR method.E.senticosus saponins content was measured by spectrophotometry method.Result: When wound method back meeting P116-1a and P116-1b after 30 d,the expression content of SS improved significantly(P<0.05),however the back meeting of P109-4 and P312-1 didn’t change the expression of SS.After that SS expression showed reduction-equality-reduction varying trend.Thirty days after back meeting P312-1,the expression content of SE improved significantly(P<0.05).Ninty days after back meeting P116-1b and P312-1,the expression content of SE improved significantly to 130%,161%,respectively(P<0.05).After 120 d,back meeting four endophytic fungi,the expression of SE were significantly higher than the control(P<0.05).Back meeting four endophytic fungi form 60 d to 120 d,the expression of bAS was significantly higher than the control(P<0.05).The back meeting four endophytic fungi improved E.senticosus saponins content significantly(P<0.05).Conclusion: Endophytic fungi P116-1a,P116-1b,P109-4 and P312-1 significantly effected the expression of key enzyme genes SS,SE and bAS and then affected E.senticosus saponins content.Among the genes,bAS was key target gene.