猪传染性胃肠炎病毒S蛋白基因的克隆与表达

根据GenBank中登录的猪传染性胃肠炎病毒(TGEV)TS株全基因的核苷酸序列,设计1对引物,采用RT-PCR技术扩增出一条含S蛋白A、D位点的S基因,大小约980bp。将其插入Simple-T克隆载体中,进行PCR、酶切鉴定及序列测定。将阳性基因插入表达载体pET-28a(+)中,转入Rossetta(DE3)感受态细胞,经诱导表达后,表达产物用SDS-PAGE和Western-blot进行分析。结果显示,表达产物的分子质量约为33.7ku,主要以包涵体的形式存在。Western-blot分析表明,表达的S蛋白能被兔源抗TGEV阳性血清所识别,说明表达产物具有反应原性。

Cloning and expression of S protein gene of swine transmissible gastroenteritis virus

According to the complete genome sequence of swine transmissible gastroenteritis virus(TGEV) TS strain in GenBank,gene of 980 bp in size for S protein with loci A and D was amplified by RT-PCR,cloned into the Simple-T vector. The constructed plasmid Simple-T-S was confirmed by restriction enzyme digestion and sequencing. The gene were then cloned into the expression vector pET-28a(+),and transfered into Rossetta(DE3) competent cells. The protein was expressed with the induction of IPTG and analyzed by SDS-P...