猪传染性胃肠炎病毒S基因B和C抗原位点片段在杆状病毒中的表达

Expression of B and C antigenic sites of TGEV spike gene in baculovirus expression system

应用RT-PCR方法扩增了猪传染性胃肠炎病毒(TGEV)TH-98株S基因5′端含有B、C抗原位点的片段。将该片段亚克隆至杆状病毒转移载体pMelBac B中。重组质粒经纯化,与线性化的杆状病毒共转染Sf9昆虫细胞,经4轮蚀斑筛选,获得了纯化的重组病毒,最终实现了在昆虫细胞内的蛋白表达。经Dot-ELISA进行抗原性分析,结果表明重组蛋白具有抗原性。本研究猪传染性胃肠炎血清学诊断方法的建立提供了必要的物质基础。

A pair of primers was designed according to the sequence published in GenBank and used to amplify the fragment containing the antigenic sites B and C of transmissible gastroenteritis virus of swine(TGEV) spike gene by RT-PCR.The PCR product was subcloned to the transfering vector pMelBac B,thus producing the recombinant pMelBac-TF.Co-transfections of Sf9 insect cells with pMelBac-TF and baculovirus linear DNA(Bac-N-Blue DNA) were performed.After purification for four times by plaque assay,a recombinant baculovirus was obtained.The antigenicity of recombinant protein was demonstrated by Dot-ELISA.This research lays a foundation for the serological diagnosis of transmissible gastroenteritis(TGE).