文档介绍：实时荧光定量PCR检测金黄色葡萄球菌femA表达水平
【摘要】目的探讨实时荧光定量PCR检测金黄色葡萄球菌femA表达的方法。方法以BB270femA基因表达为标准,建立实时荧光定量PCR方法,对实验菌株femA表达进行相对定量。结果建立的实时荧光定量PCR方法所得结果标准曲线线性关系好,熔解峰单一。随MIC变化,金黄色葡萄球菌femA表达水平也随之变化。结论用实时荧光定量PCR方法研究金黄色葡萄球菌femA表达量结果可靠、敏感、重复性好。
【关键词】金黄色葡萄球菌;femA基因;实时荧光定量PCR
Abstract: Objective To establish the approach of real-time fluorescent quantitative PCR to detect femA gene in us aureus strains. Methods Real-time fluorescent quantitative PCR was performed to quantify the expression of femA gene of 15 clinical stains, pared with BB270. Results The standard curve by real-time fluorescent quantitative PCR showed good linearity between the amount of femA RNA and cycle threshold and the melting curve had a single peak. The variation of expression level of femA was in accordance with MIC of strains. Conclusions The real-time fluorescent quantitative PCR in the detection of femA expression is accurate, sensitive and repeatable.
Key words: us aureus; femA gene; real-time fl
uorescent quantitative PCR
实时荧光定量PCR(real-time fluorescence quantitative PCR,real-time Q-PCR)由于其具有特异性强、灵敏度高、重复性好、定量准确、速度快等优点,目前已经被广泛应用于基础科学研究、临床诊断、疾病研究及药物研发等领域中,其中最主要的应用集中在以下几个方面:DNA或RNA的绝对/相对定量分析,基因表达差异分析,基因分型。
耐甲氧西林的金黄色葡萄球菌(methicillin-resistant us aureus,MRSA)是全球感染性疾病领域研究热点之一。已知femA是MRSA耐药表达的重要辅助基因,与MRSA高水平耐药成正相关[1-3],但还未见研究报道两者确切关系。本实验利用近年来兴起的实时荧光定量PCR检测金黄色葡萄球菌femA基因表达水平,进一步明确femA在MRSA耐药表达中的意义。同时建立实时荧光定量PCR用于金黄色葡萄球