刺五加鲨烯合酶和鲨烯环氧酶基因单核苷酸多态性及其与总皂苷量的相关性研究

Single nucleotide polymorphism of squalene synthase and squalene epoxidase genes and their correlation with content of saponins in Eleutherococcus senticosus

目的筛选刺五加鲨烯合酶(SS)和鲨烯环氧酶(SE)基因存在的单核苷酸多态性(SNP)位点,分析各SNP位点与刺五加总皂苷量的相关性。方法采用分光光度法测定刺五加总皂苷的量,并划分为具有显著差异(P<0.01)的高、低量组;RT-PCR法扩增刺五加SS、SE基因,根据测序结果筛选SNP位点,利用R×C表的χ2检验分析相关关系。结果刺五加SS基因存在6处SNP,其中704 bp位点为错义突变,其他位点为同义突变,各位点均不与刺五加总皂苷量显著相关。SE基因存在9处SNP,其中导致错义突变的164、199、227、232 bp位点及同义突变的279、285 bp位点与刺五加总皂苷量显著相关(P<0.05),其他位点为同义突变,与刺五加总皂苷量间无显著相关性。结论刺五加SS和SE基因存在SNP,SE基因164～285 bp位点的AGAACG与刺五加总皂苷高量组显著相关,TAGTTC与低量组显著相关。

Objective To screen the single nucleotide polymorphism（SNP） sites of squalene synthase（SS） gene and squalene epoxidase（SE） gene in Eleutherococcus senticosus and to analyze the correlation between each SNP site of SS and SE genes and the content of the total saponins in E.senticosus.Methods The contents of the total saponins in E.senticosus were detected by spectrophotometric method and divided into high-and low-content groups with significant difference（P 0.01）.RT-PCR method was developed to amplify SS and SE genes.SNP sites were screened according to sequencing result.The correlation was observed by R × C χ2test and analysis.Results Six SNP sites existed in SS gene of E.senticosus,among which 704 bp site led to missense mutation and other SNP sites were synonymous mutations.No SNP site in SS had significant correlation with the content of the total saponins in E.senticosus.There were nine SNP sites in SE gene.Sites at 164,199,227,and 232 bp were missense mutation while at 279 and 285 bp were synonymous mutations,and all of them had the significant correlation with the content of the total saponins in E.senticosus（P 0.05）.Other SNP sites of SE gene were synonymous mutations and had no significant correlation with the content of the total saponins in E.senticosus.Conclusion There are SNP sites in SS and SE genes of E.senticosus.AGAACG located in 164—285 bp of SE gene could significantly correlate with the high-content group of the total saponins in E.senticosus.TAGTTC located in the same sites could significantly correlate with the low-content group of the total saponins in E.senticosus.