猪圆环病毒2型截短衣壳蛋白的表达及抗原性鉴定
Expression and antigenicity characterization for truncated capsid protein of porcine circovirus type 2.
作者信息
Lou Zhongzi, Li Xuerui, Li Zhiyong, Yin Xiangping, Li Baoyu, Lan Xi, Yang Bin, Zhang Yun, Liu Jixing
机构信息
State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, China.
出版信息
Can J Vet Res. 2011 Jan;75(1):61-4.
Three pairs of specific primers were designed to amplify F2-1, F2-2, and XF2-2 truncated capsid protein genes of porcine circovirus type 2 (PCV-2). Amplified sequences were subcloned to pET-32a(+) vectors and expressed in Rosetta (DE3) Escherichia coli by induction of isopropy-β-D-thiogalactoside (IPTG). All of the fusion proteins had positive reactions to PCV-2 antiserum and His-XF2-2 showed the best reactivity. Proteins were used to immunize BALB/c mice to produce monoclonal antibodies (mAbs), and 7 mAbs were selected. Capsid protein N-terminal parts 55 to 96 amino acid (aa), 97 to 141 aa, and 143 to 211 aa were confirmed as binding regions of the 7 mAbs. Reactivity between His-XF2-2 and the 7 mAbs was detected, FmAb-8 showed the best reactivity. The dominant B-cell epitope was located at 97 to 141 aa. The PEPSCAN indicated that the P122-136 peptide contained the dominant B-cell epitope.
设计了三对特异性引物，用于扩增猪圆环病毒2型（PCV-2）的F2-1、F2-2和XF2-2截短衣壳蛋白基因。将扩增的序列亚克隆到pET-32a(+)载体中，并通过异丙基-β-D-硫代半乳糖苷（IPTG）诱导在Rosetta (DE3)大肠杆菌中表达。所有融合蛋白对PCV-2抗血清均有阳性反应，His-XF2-2的反应性最佳。用这些蛋白免疫BALB/c小鼠以产生单克隆抗体（mAb），并筛选出7种mAb。衣壳蛋白N端第55至96个氨基酸（aa）、第97至141个aa和第143至211个aa被确认为这7种mAb的结合区域。检测了His-XF2-2与这7种mAb之间的反应性，FmAb-8的反应性最佳。主要B细胞表位位于第97至141个aa处。PEPSCAN表明P122-136肽段包含主要B细胞表位。
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