卡氏肺孢子虫MSG抗原基因片段真核表达质粒构建与表达

CONSTRUCTION AND EXPRESSION OF EUKARYOTIC EXPRESSION PLASMIDS OF PNEUMOCYSTIS CARINII MSG ANTIGEN GENE FRAGMENT

[目的]构建大鼠源卡氏肺孢子虫MSG抗原基因片段真核表达质粒,进行表达。[方法]以卡氏肺孢子虫DNA为模板,应用PCR扩增MSG基因片段,连接至pGEM-T Easy载体,再克隆至PCDNA3.1(+)载体。构建的重组表达质粒经酶切、PCR鉴定后转染至COS-7细胞,并进行大量增殖。RT-PCR验证转染基因的表达。[结果]重组表达质粒PCDNA3.1(+)/MSG经酶切及PCR鉴定结果表明构建成功。RT-PCR结果显示MSG基因片段成功转染至COS-7细胞,并在其中进行基因表达。[结论]成功构建了PCDNA3.1(+)/MSG重组质粒,并在COS-7细胞中表达,为进一步进行核酸疫苗的研究打下基础。

[Objective] To construct eukaryotic expression plasmids of Pneumocystis carinii （PC） MSG antigen gene fragment, and proceed on expression. [Methods] The PC MSG antigen gene fragment was amplified from lung tissue of rats infected by PC and cloned into pGEM-T Easy vector. After identification, the fragment was inserted into eukaryotic expression vector, and cloned to PCDNA3.1 （＋） cartier. The recombinant plasmid was screened and identified by PCR and restriction analysis to transfect to COS-7 cell, and greatly proliferation. The RT-PCR verified the expression of transfection gene. [ Resuits ] The recombinant plasmid PCDNA3.1 （＋） /MSG was constructed successfully by the verification of PCR and restriction analysis. Tile result of RT-PCR confirmed that the MSG gene fragment was transfected into COS-7 cells, and the gene was expressed into the cells. [ Conclusion] Construction and expression of eukaryotic expression plasmid PCDNA3.1 （＋） /MSG has been constructed successfully and expressed in COS-7 ceils, this study provides basis for the further gene engineering vaccine study.