SARS-COV S蛋白在大肠杆菌和家蚕中的表达

将SARS-CoV S蛋白基因的部分序列(accession number为AY304495的22 908-23 542 nt区域),克隆到表达载体pET28 a(+)中,并在大肠杆菌BL21中诱导表达。SDS-PAGE、W estern b lotting分析表明,重组S蛋白的分子量约为27 kD,与理论分子量相符。将S蛋白基因的部分序列克隆入杆状病毒转移载体pBacPAK-H is后,与线性化家蚕杆状病毒BmBacPAK-6共转染BmN细胞,经空斑筛选获得重组杆状病毒BmBac-S,并将其在细胞和家蚕中进行表达。SDS-PAGE、W estern b lotting分析表明,表达产物的分子量约30 kD,用金属螯合亲和层析纯化得到家蚕细胞表达的重组S蛋白。

Expression of SARS Coronavirus Spike Protein in E. coli and Silkworm Bombyx mori

The part sequence(base pairs from(22 908) to(23 542) in SARS-CoV genome sequence whose accession number is AY304495) of spike protein gene of SARS Coronavirus was cloned into the expression vector pET28a(+).SDS-PAGE and western blotting demonstrated that in E.coli BL21 recombinant S protein gene was expressed with the molecular weight about 27 kD of identity with its theoretical molecular weight.Also this part sequence of spike protein gene was inserted into baculovirus transfer vector pBacPAK-His,then cotransfected with liner silkworm baculovirus BmBacPAK-6 DNA in BmN cells to construct the recombinant baculovirus BmBac-S.Plaque assay was taken to screen it.SDS-PAGE and western blotting analysis performed that the molecular weight of recombinant S protein was about 30 kD in silkworm.The pure recombinant S protein expressed by baculovirus could be easily obtained through one-step metal chelate affinity chromatographer.