黄芪中杀菌抑菌单体分子的筛选及差异表达基因检测  

Screening of bactericidal and bacteriostatic monomer molecules in Astragali Raidx and detection of differentially expressed genes

作　　者：赵欣如 周润龙 明文卓 王璐 张宏飞 张稳稳 李诚博 ZHAO Xinru;ZHOU Runlong;MING Wenzhuo;WANG Lu;ZHANG Hongfei;ZHANG Wenwen;LI Chengbo(Binzhou Medical University,Yantai 264003,Shandong,P.R.China)

机构地区：[1]滨州医学院,山东烟台264003

出　　处：《滨州医学院学报》2024年第4期248-252,共5页Journal of Binzhou Medical University

基　　金：山东省高等学校青创科技支持计划(2020KJK003,2022KJ280);烟台市科技创新发展计划(2023JCYJ079);大学生创新创业训练计划(S202210440001)。

摘　　要：目的筛选黄芪中有效抑制金黄色葡萄球菌(SA)的单体分子,筛选有效单体分子作用SA后的差异表达基因。方法针对黄芪中6种单体分子(黄腐酚、芒柄花黄素、柚皮素、环黄芪醇、黄芪皂苷Ⅰ、黄芪甲苷),使用酶标仪检测法进行SA的体外抑菌实验(MIC),筛选出对SA有良好抑制作用的单体分子。使用梯度稀释点样法针对有良好抑制作用的单体分子进行体外杀菌实验(MBC),验证其杀灭SA的效果,并通过设置不同的浓度组确定其最佳杀菌浓度。将有效抗菌的单体分子作用后的SA与未加药的SA收样,利用RNA-seq技术进行转录组分析,比对出有效单体分子作用后的差异表达基因。结果MIC实验筛选出对SA有抑制作用的3种单体分子,其中黄腐酚抑制效果最佳,芒柄花黄素有较好的抑制效果,柚皮素有抑制作用但效果不佳,其他3种单体对SA无抑制作用。MBC实验表明黄腐酚对SA具有良好杀菌效果,且最佳杀菌浓度为12.5μg·mL^(-1)。经转录组测序,与未加黄腐酚组相比,加入黄腐酚的组别共检测到了63个显著性差异表达基因,其中有10个上调基因,53个下调基因,并且大部分基因都和核糖体蛋白合成相关。结论黄腐酚对SA具有良好的杀菌抑菌作用,通过转录组测序发现黄腐酚可能是通过抑制核糖体内的蛋白质合成,使SA的蛋白质合成过程受阻,进而导致SA死亡。Objective To screen effective monomeric molecules against Staphylococcus aureus(SA)in Astragali Raidx,and to screen differentially expressed genes following the action of effective monomeric molecules.Methods In vitro inhibition assay(MIC)of SA was conducted for 6 monomer molecules from Astragali Raidx,and the monomer molecules with good inhibition effect on SA were screened.In vitro bactericidal assay(MBC)was conducted to verify the sterilization function of the monomer molecules with good inhibitory effect on SA,and the optimal bactericidal concentration was determined by setting different concentration groups.The samples of SA after the action of the effective bactericidal inhibitory monomer molecules were collected with the unspiked SA,and the transcriptome analysis was performed by RNA-seq technology to compare the differentially expressed genes after the action of the effective monomer molecules.Results The MIC experiment screened 3 monomer molecules with inhibitory effect on SA,among which Xanthohumol had the best inhibition effect,Formononetin had good inhibition effect,Naringenin had inhibition effect but not good effect,and the other 3 monomeric molecules without good inhibition effect on SA were screened.MBC experiment verified that Xanthohumol had good bactericidal effect on SA,and the best bactericidal concentration was 12.5μg·mL^(-1).After transcriptome sequencing,a total of 63 significantly differentially expressed genes were detected in the group with Xanthohumol added compared to the group without Xanthohumol,including 10 upregulated genes and 53 downregulated genes.Most of the genes were related to ribosomal protein synthesis.Conclusion Xanthohumol had a good bactericidal and bacteriostatic effect on SA.Through transcriptome sequencing,it was found that Xanthohumol might inhibit the biosynthesis of ribosomes,thus hindering the protein synthesis process of SA and leading to SA death.

关 键 词：黄芪 金黄色葡萄球菌 黄腐酚 抗菌活性 蛋白质合成 

分 类 号：R286[医药卫生—中药学]