摘要
将猪瘟兔化弱毒株Erns基因亚克隆到原核表达载体pGEX-6p-1中,经PCR和双酶切鉴定以及序列分析,表明已成功构建了重组质粒。将重组质粒转化到大肠埃希氏菌BL21(DE3)中进行表达,经SDS-PAGE和Western-Blot检测,结果表明,融合表达蛋白的分子量约为55kD,目的蛋白表达量占菌体总蛋白的45%。表达蛋白与兔抗CSFV多抗发生特异性反应,具有良好的反应原性。利用GST亲和层析柱对Erns融合蛋白进行了纯化。
The full lengh of E^rns gene was subcloned into the prokaryotic expression vector pGEX - 6p - 1. The results of PCR amplification, restriction enzyme digestion and DNA sequencing showed that the recombinant plasmid was obtained. Then it was transformed into E. coli BL21（DE3）competent cells for expression. The expressed protein was identified by SDS - PAGE and Western - Blot, which showed the molecular weight of E^rns fusion protein was approximately 55KD. E. coli BL21 （DE3） harboring the recombinant plasmid pGEX- 6p- 1 -E^rns expresses the protein at high level （45%）. The expressed protein was specifically recognized by rabbit anti-CSFV polyclonal antibodies. E^rns protein was purified by GST - chelating chromatography. This may provide tools for further development of diagnostic reagent for detecting CSF antibodies.
出处
《河南农业科学》
CSCD
北大核心
2009年第12期121-123,127,共4页
Journal of Henan Agricultural Sciences
基金
国家"863"计划项目(2007AA100606)
关键词
猪瘟兔化弱毒
Erns基因
原核表达
纯化
猪瘟
Hog cholera lapinised virus （HCLV）
E^rns gene
Prokaryotic expression
Purification
CSF
