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中文摘要：
目的利用杆状病毒系统表达hGITRLaa52-177蛋白。方法PCR扩增获得hGITRL胞外段(hGITRLaa52-177)基因,克隆入pFastBacHTa转移载体,在E.coliDH10Bac内通过转座子Tn7的介导,得到重组杆粒,转染Tn细胞表达目的蛋白。通过SDS-PAGE电泳及Westernblotting鉴定目的蛋白的表达。结果重组载体pFastBacHTa-hGITRLaa52-177以及重组杆粒Bacmid-hGITR-Laa52-177构建正确,在Tn细胞内成功表达6×His-hGITRLaa52-177重组蛋白,SDS-PAGE电泳及Westernblotting显示重组hGITRLaa52-177蛋白相对分子质量约为Mr18000。结论杆状病毒-昆虫表达系统获得hGITRLaa52-177蛋白,为进一步从蛋白质水平研究hGITR/hGITRL对机体免疫调节的影响奠定了基础。
英文摘要：
Objective To produce the extracellular region of hGITRL in eukaryotic expression system.Methods Bac-to-Bac baculovirus expression system was employed in the present study.The hGITRL extracellular region gene was amplified by PCR from the full-length pMD18-hGITRL-T vector and subcloned into the pFastBacHTa plasmid,then the latter was transformed into E.coli DH10Bac competent cells to construct a recombinant baculovirus and to infect Tn cells.The interest protein was identified by SDS-PAGE,Western blot and in...
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