结核分支杆菌Rv3117基因的克隆与原核表达
Progress in Veterinary Medicine（2015）
Abstract
以结核分支杆菌H37Rv株染色体DNA为模板,应用Rv3117基因特异性引物对该基因进行PCR扩增,获得约800 bp的DNA片段.将PCR纯化产物克隆至pMD-18T Vector中,构建出重组质粒pMD-18T-Rv3117.以EcoR Ⅰ和SamⅠ双酶切pMD-18T-Rv3117重组载体和pGEX-4T-1原核表达载体,并将纯化的Rv3117基因亚克隆至pGEX-4T-1中,构建出原核重组表达质粒pGEX-4T-1-Rv3117.将pGEX-4T-1-Rv3117重组质粒转化至感受态E.coli BL21中,经IPTG诱导和SDS-PAGE分析,可见约30ku目的蛋白带.Western blot分析结果显示,该蛋白能与抗结核分支杆菌阳性血清发生特异性反应.研究结果为结核病临床诊断抗原的筛选奠定了基础.
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