小反刍兽疫病毒H基因在杆状病毒中的表达

从小反刍兽疫病毒全长cDNA中对血凝素蛋白H基因进行特异扩增,回收PCR产物分别连接于T载体酶切及测序分析后,将其亚克隆至真核表达转移载体pFastBacHT,经重组筛选获得杆状病毒重组质粒。重组质粒转染sf9细胞后连续传3～4代,分别收获细胞上清及沉淀用于SDS-PAGE及Western blot对重组H蛋白的检测,用抗His蛋白单抗在细胞中检测到H标签蛋白,单抗在细胞及上清中检测到大小为46 ku的融合蛋白。

EXPRESSION OF H GENE OF PESTE DES PETITS RUMINANTS VIRUS IN BACULOVIRUS

The H gene fragment of Peste des petits ruminants virns strain N75/1 was amplified from cDNA library by PCR.Purified PCR products were cloned into pMD19-T vector.After identification with enzyme digestion and sequence analysis,H gene was subcloned into transfer vector pFastBacHT.The baculoviruses containing recombinant plasmid were selected and then inoculated into Sf9 cells.Supernatants and cell pellets were harvested from infected Sf9 cells after 3-4 passages and subject to SDS-PAGE and Western blot.The H protein with molecular mass of 46ku was detected with anti-His tag protein in both cell pellets and supernatants.