中华蜜蜂囊状幼虫病病毒非结构蛋白抗体的制备及其应用

Antiserum preparation and application of polychonal antibodies against non-structural protein of Chinese sacbrood virus

应用Gateway重组技术构建CSBV非结构蛋白,RNA依赖的RNA聚合酶(RNA-dependent RNA polymerase,RdRp)的原核表达载体,将重组表达载体转化大肠杆菌Rosetta,经异丙基硫代-β-D-半乳糖苷(Isopropylβ-D-1-Thiogalactopyranoside,IPTG)诱导获得RdRp基因原核表达蛋白.以重组蛋白为抗原免疫新西兰兔制备相应的多克隆抗体,Western blot检测表明,该抗体能与RdRp重组蛋白和感病中华蜜蜂幼虫蛋白特异性结合,说明获得的抗体特异性高.利用制备的抗体进行免疫荧光标记,发现其能特异地定位在感病中华蜜蜂血淋巴细胞内.

The expression plasmids of non-structural protein, RNA-dependent RNA polymerase （RdRp） of CSBV were constructed by the technology of Gateway and then transformed into E. eoli strain Rosetta. Recombinant protein was obtained through Isopropyl 13-D-1-Thiogalactopyranoside （IPTG） induction. The recombinant protein was used to immunize rabbits for preparation of polyclonal antibodies. The analysis of Western blot indicated that the prepared antibodies could specially bind with recombined RdRp protein and protein of Ap/s cerana cerana infected by CSBV, respectively. It was observed that CSBV-RdRp protein was specially located in the haemocytes of A. c. cerana infected by CSBV using immunofluorescence labeling.