摘要
针对猪链球菌2型(SS2)溶菌酶释放蛋白(MRP)和纤维蛋白原结合蛋白(FBP)抗原性较强的基因片段,分别设计了1对引物,用PCR扩增出了mrp基因和fbp基因的片段。利用这些片段分别构建了原核表达载体pET32a-mrp、pET32a-fbp和pET32a-mrp-fbp。将这些表达载体分别转化大肠杆菌BL21(DE3),在IPTG诱导下进行了表达。SDS-PAGE分析表明,表达的MRP、FBP、MRP-FBP的分子质量分别为47、52、80 ku。Western-blot分析表明,重组蛋白MRP-FBP可被SS2阳性血清所识别。免疫保护试验表明,串联表达的融合蛋白MRP-FBP对BALB/c小鼠的保护率达80%,是目前筛选到的制备SS2亚单位疫苗保护力最高的短片段串联免疫原。
Two pairs of primers were designed to amplify the gene fragments encoding muramidasereleased protein（MRP） and fibrinogen-binding protein（FBP） of Streptococcus suis type 2（SS2）. The recombinant expression plasmids pET32a-mrp, pET32a-fbp and pET32a-mrp-fbp were respectively constructed with the PCR-amplified fragments of mrp and fbp genes, and then transformed into Escherichia coli BL21（DE3） induced by IPTG. SDS-PAGE analysis showed that the expressed MRP,FBP and MRP-FBP was 47 ku,52 ku and 80 ku in molecular mass respectively. Western-blot analysis showed that MRP-FBP could be recognized by the serum against SS2 whole cell. Adhesin vaccines were prepared respectively with MRP,FBP and MRP FBP as antigen. To determine protective efficacy of the three vaccines,the vaccine with MRP-FBP could protect 80% of the mice to survive by the test of mice vaccination. It was concluded that the recombinant MRP-FBP was the best candidate antigen for the development of subunit vaccines against SS2 up to date.
出处
《中国兽医科学》
CAS
CSCD
北大核心
2009年第6期522-526,共5页
Chinese Veterinary Science
基金
国家自然科学基金项目(30571389)
甘肃省农业生物技术研究与应用开发项目(GNSW-2008-09)
中国博士后科学基金项目(20080430788)
甘肃省自然科学基金项目(3ZS051-A25-060)
