摘要
将RT-PCR扩增的猪传染性胃肠炎病毒(TGEV)S基因A抗原位点目的片段克隆、双酶切后连接于表达载体,经酶切、PCR和测序鉴定的阳性重组质粒转化BL21(DE3),用IPTG诱导,进行SDS-PAGE和Western-blotting分析,确定目的蛋白的原核表达情况和免疫特异性。结果显示,目的片段的大小为534bp,序列分析表明,该基因与其他TGEV相应基因具有很高的同源性;Western-blotting检测可见分子质量约43 ku的融合蛋白条带,表明,该蛋白具有良好的反应原性。
Antigenic site A sequence amplified by RT-PCR and cloned into th of S gene of swine transmissible gastroenteritis virus（TGEV） was e prokaryotic expression vector after digestion with restriction enzyme. The positive recombinants identified by enzymatic digestion, PCR and sequencing was transformed into Escherichia coli BL21 and the recombinant transformant was induced with IPTG. The prokaryotic expression and the immunologic specificity of the expressed protein were analyzed by SDS-PAGE and Western-blotting. The results showed that the target gene was 534 bp in length and had a relatively high homology with the corresponding gene from other TGEV strains. The expressed fusion protein was 43 ku in size,and had good reactogenicity as revealed by Western-blotting.
出处
《中国兽医科学》
CAS
CSCD
北大核心
2007年第10期845-849,共5页
Chinese Veterinary Science
基金
农业应用新技术北京市重点实验室开放课题资助项目(KF2004-04)
关键词
猪传染性胃肠炎病毒
S基因A抗原位点
克隆
原核表达
swine transmissible gastroenteritis virus
antigenic site A of S gene
cloning
prokaryotic expression
