传染性法氏囊病病毒VP2基因在昆虫细胞中的表达

Expression of VP2 Gene of Infectious Bursal Disease Virus in Insect Cells

本研究将鸡传染性法氏囊病病毒超强毒（very virulent infectious bursal disease virus,vvIBDV）JM-1/10株VP2基因克隆至pcDNA-3.1（＋）启动子CMV之后,随后将CMV-VP2基因序列一同克隆入杆状病毒表达系统的质粒 pFastBacTM Daul中构建了pFast-CMV-VP2。将pFast-CMV-VP2转化Escherichia coli DH10Bac感受态细胞,筛选出重组质粒Bacmid-CMV-VP2。用 Bacmid-CMV-VP2转染Sf9昆虫细胞,获得了重组杆状病毒vBac-CMV-VP2。将该重组杆状病毒转导BHK-21细胞,48~72 h后经间接免疫荧光试验（IFA）检测到VP2蛋白具有特异性荧光;样品经Western blotting分析,结果显示目的蛋白得到表达。结果表明,本试验制备的重组 vBac-CMV-VP2 既能在昆虫细胞中表达,也可在哺乳动物细胞中表达。

The VP2 gene of very virulent infectious bursal disease virus （vvIBDV） JM-1/10 strain was placed behind the CMV promoter of pcDNA-3.1（＋）.The CMV-VP2 gene was cloned into the baculovirus expression system pFastBacTM Dual to build pFast-CMV-VP2.Transformed it to Escherichia coli DH10Bac.Recombinant baculovirus expression plasmid Bacmid-CMV-VP2 was screened.Bacmid-CMV-VP2 was used to be transfected into Sf9 insect cells to get the recombinant baculovirus vBac-CMV-VP2 and transduced into BHK-21 cells.After 48 to 72 hours,indirect immunofluorescence assay （IFA） was conducted to detect specific fluorescence.Western blotting analysis result showed target protein was expressed.The prepared recombinant vBac-CMV-VP2 could not only be expressed in insect cells, but also expressed in mammalian cells.