摘要
利用PCR方法扩增乳源致病性金黄色葡萄球菌nEBPS全基因,将其定向克隆至原核表达载体pET30a(+)中,鉴定正确后,转化BL21表达菌,经IPTG诱导获得了以可溶性表达的重组蛋白。通过Ni NTA Purification System纯化重组蛋白,纯化蛋白质量浓度为2.16g/L。纯化蛋白经免疫印迹检测显示重组蛋白能够被牛源金黄色葡萄球菌阳性血清识别,具有良好的反应性。将纯化的重组蛋白免疫新西兰白兔制备多克隆抗体,间接ELISA测定抗体效价为1∶25 600,凝集试验测定抗体效价为1∶128。
To expression of N-terminal elastin binding proteins of Staphylococcus aureus （nEBPS） and prepare polyclonal antibody, the nEBPS was amplified by PCR and directionally cloned into the pET30a vector. After characterization, the recombinant plasmid was transformed into E. coli BL21（DE3）,the recombinant nEBPS protein was expressed in soluble body form after induction with IPTG. The concentration of the purified recombinant protein was 2.16 g/L. Western blotting showed that purified protein could react specifically to Staphylococcus aureus positive serum. The polyclonal antibodies to the nEBPS protein was prepared by inoculating rabbits with purified re combinant protein. The polyclonal antibodies was generated with antibody titres of 1 ： 25 600 in ELISA and 1 ： 128 in agglutination test. The recombinant protein and the polyclonal antibody ob- tained in this study could be used for detection Staphylococcus aureus,and could be used to fur ther study the structure,function and epitope mapping of nEBPS.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2013年第11期1674-1678,共5页
Chinese Journal of Veterinary Science
基金
国家自然科学基金资助项目(31060352)
内蒙古自治区科技厅科技创新引导计划资助项目(20111802[2012])
关键词
金黄色葡萄球菌
弹性纤维结合蛋白N端蛋白
原核表达
多克隆抗体
Stapkylococcus aureus
N-terminal elastin binding proteins of Staphylococcus aureus
prokaryotic expression
polyclonal antibody
