鸡β-防御素-9的融合表达及其多克隆抗体的制备与鉴定

摘要

为制备鸡β-防御素-9(AvBD9)多克隆抗体,本试验将AvBD9基因插入pGEX-6P-1载体,构建重组基因pGEX6P-AvBD9大肠杆菌表达质粒.将通过切胶纯化获得的表达产物包涵体谷胱甘肽转移酶-AvBD9(GST-AvBD9)融合蛋白免疫新西兰大白兔,制备抗AvBD9血清,并检测抗体效价.结果表明,经间接酶联免疫分析(ELISA)及蛋白质免疫印迹(Western blot)方法证实,获得的融合蛋白免疫新西兰大白兔得到了特异性的多克隆抗体,抗体效价为1:38 400.%To prepare polyclonal antibody against chicken β-defensin-9 (AvBD9) , AvBD9 gene was cloned into the prokaryotic expression vector pGEX-6P-1. The recombinant expression plasmid pGEX-6P-AvBD9 was constructed and then transformed into Escherichia coli (E. coli) to express AvBD9. The glutathione transfer-ase-AvBD9 (GST-AvBD9) fusion protein was expressed in E. coli as an inclusion body. The fusion protein purified from the PAGE gel was used to immunize New Zealand white rabbit. The results showed that as detected by indirect ELISA and Western blot, polyclonal antibody against AvBD9 with the titer of 1:38 400 was obtained from the immunized rabbits.