猪链球菌2型MRP与FBPS部分片段串联基因的原核表达

Cloning and Expression of a Fusion Protein of muramidase-released protein and fibronectin and fibrinogen-binding protein of Streptococcus suis type 2

根据猪链球菌2型(SS2)溶菌酶释放蛋白(MRP)和纤维蛋白原结合蛋白(FBPS)的基因序列,选取可能含有抗原决定簇的两片段,各设计合成一对引物,使之含有共同的酶切位点。通过T4DNA连接酶串联两片段,构建原核表达载体pET32a-mrp-fbps,重组质粒转化入大肠杆菌BL21(DE3),经不同时间、不同温度,不同浓度IPTG诱导表达,经SDS-PAGE电泳,表达约为80kD的融合蛋白。Westernblot结果表明重组蛋白可被SS2阳性血清所识别。证实串联表达的融合蛋白具有良好的免疫原性,是重要的保护性抗原,本试验为进一步研究猪链球菌亚单位疫苗奠定了基础。

to the gene sequence of Streptococcus suis type 2 muramidase-released protein （MRP） and fibronectin and fibrinogen-binding protein （FBPS）, two fragments probably containing epitopes were selected, and a pair of primers having common restriction sites were designed. The two fragments were linked to construct the prokaryotic expression vector pET32a-mrp-fbps. And the re-assembled plasmid was then transferred into E. coli BL21 （DE3）.SDS-PAGE electrophoresis revealed that about 80kD fusion protein was expressed through induced expression, for different time, and at different temperature and concentration. The Western blot result demonstrated that the recombinant protein can be recognized by SS2 positive serum and fusion protein in series expression is a sort of important protective antigen. This experiment provided the foundation for further research of swine streptococcus subunit vaccine.