To express the actin depolymerizing factor (ADF) of Eimeria stiedae in HeLa cells, the ADF gene was amplification from the E. Stiedae total RNA by RT-PCR and cloned into pVAX-1. The resultant recombinant plasmid was transfected into HeLa cells with FuGENE. The results indicated that the ADF gene of E. Stiedae was expressed in HeLa cells detected by RT-PCR and western blot, which would be useful for evaluating immunogenicity of ADF gene as a DNA vaccine against E. Stiedae.%为真核表达斯氏艾美尔球虫(Eimeria stiedae)肌动蛋白解聚因子(ADF)基因,本研究采用RT-PCR技术扩增ADF基因,构建pVAX- 1-ADF真核重组质粒,并转染HeLa细胞.采用RT-PCR方法检测ADF mRNA转录水平、western blot检测ADF蛋白表达水平.琼脂糖凝胶电泳结果表明PCR扩增出一条大小为350 bp的特异性片段,与预期值相符.Western blot检测显示,转染pVAX- 1-ADF的HeLa细胞在大约13ku处出现特异性表达条带,与预期值相符.本研究构建了E.stiedae ADF基因真核表达质粒pVAX- 1-ADF,并且在体外真核细胞中表达,为进一步寻找防治E.stiedae的疫苗候选基因奠定了基础.
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