H9N2禽流感病毒HA2基因重组杆状病毒的表达

从pMD18-T-HA阳性质粒扩增了H9N2亚型禽流感病毒的HA2基因,将扩增到的 HA2基因克隆至昆虫杆状病毒转移载体pBlueBacHis2A中。将其与杆状病毒共转染于Sf9昆虫细胞,经蚀斑筛选纯化重组杆状病毒,用其感染Sf9昆虫细胞,并优化表达条件。SDS-PAGE和 Western-blotting分析表明,表达产物的分子质量约为27 ku。Dot-ELISA分析表明,表达的HA2 融合蛋白可与鸡抗H9N2亚型血清发生特异性反应,而与H5和H7亚型抗血清间无交叉反应。

Recombinant baculovirus expression of HA2 gene of avian influenza virus H9N2 subtype

Using a pair of specific primers designed according to the pMD18-T-HA, the HA2 gene of avian influenza virus H9N2 subtype was amplified by PCR. The PCR product was cloned into pBlueBa-cHis2A to obtain an eukaryotic recombinant plasmid pBlueBacHis2A-HA2, and then Sf9 cells were trans-fected with the recombinant plasmid. After plaque screening, the target HA2 gene was expressed in insect cell Sf9, and the expressed protein was 27 ku in molecular weight. Western-blotting and Dot-ELISA analysis showed that the recombinant protein had good reactivity against H9N2 subtype serum and did not have any cross-reaction with chicken H5 and H7 antiserum.