Effect of environmental factors on Listeria monocytogenes inl B gene expression
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摘要: 针对单增李斯特菌毒力基因inl B设计五对引物,从中筛选出一对特异性强的引物,利用反转录荧光定量PCR技术来考察毒力基因表达的强弱。在不同环境因素条件下培养单增李斯特菌,分别提取菌体RNA,继而合成c DNA,以管家基因16S r RNA为内参基因,利用循环阈值(Ct)的变化计算inl B基因的相对表达量。结果表明,毒力基因inl B在温度为15℃、p H为6、氯化钠浓度为1.5%~2.0%、蔗糖浓度为0.75%~1.25%时能高效表达;温度为0℃和45℃、p H为4和9、氯化钠浓度为3.5%以上以及蔗糖浓度为0%~0.25%和2.25%以上时,对毒力基因inl B的表达起明显的抑制作用;在p H为6的条件下,酸的种类对毒力基因inl B的表达影响不大。Abstract: Five pairs of primers were designed based on Listeria monocytogenes virulence genes inl B and a pair of well specific primers were selected for this study. The Listeria monocytogenes virulence factor expression was studied by real-time reverse transcriptase PCR. RNAs were extracted from Listeria monocytogenes culture solution under the different environmental factors conditions,and then c DNAs were synthesized respectively.The housekeeping gene 16 S r RNA was used as an internal standard. The changes of the Ct value could mirror the relative expression levels of inl B gene. Our data suggested that inl B gene expression was efficient when the temperature was 15 ℃,p H was 6,the Na Cl concentration was 1.5% to 2%,and the concentration of sucrose was 0.75% to 1.25%. When the temperature was 0 ℃ or 45 ℃,p H was 4 or 9,the Na Cl concentration was more than 3.5%,and the concentration of sucrose was 0% to 0.25%,or more than 2.25%,inl B gene expression was suppressed. What's more,the kinds of acid had little effect on the inl B gene when the p H was 6.
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