| 鲤春病毒血症病毒G蛋白的原核表达及多克隆抗体的制备 |
| Prokaryotic Expression and Polyclonal Antibody Preparation of G Protein of Spring Viremia of Carp Virus |
| 投稿时间：2014-03-04 修订日期：2014-03-26 |
| DOI： |
| 中文关键词:鲤春病毒血症病毒 G蛋白 原核表达 多克隆抗体 |
| 英文关键词:Spring viremia of carp virus G protein prokaryotic expression polyclonal antibody |
| 基金项目:国家自然科学基金项目（31172433，30901118）；中央高校基本科研业务费专项资金资助项目（2013PY071，2011PY121） |
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| 中文摘要: |
| 利用PCR方法扩增鲤春病毒血症病毒糖蛋白G的部分基因序列，即全基因组序列上第3094～4170位的碱基，并将其克隆至表达载体pGEX-KG中，构建重组质粒SVCV-g-KG。将重组质粒转化感受态细胞BL21，经IPTG诱导后，表达了与预期大小相符的约66 kDa的融合蛋白，可溶性分析表明该蛋白主要表达在包涵体中。将纯化的融合蛋白免疫日本大耳白兔，制备多克隆抗体。酶联免疫吸附试验检测其抗体效价可达1:256000，间接免疫荧光试验和免疫印迹试验证明其与病毒结合活性良好，特异性高；病毒孵育试验结果表明该多克隆抗体具备中和活性，能够有效阻断鲤春病毒血症病毒对细胞的感染。 |
| 英文摘要: |
| Glycoprotein G gene segment(3094～4170bp) of spring viremia of carp virus (SVCV) was amplified by PCR and cloned into the prokaryotic expression vector pGEX-KG. The recombined plasmid SVCV-g-KG was transformed into E.coli BL21 and fusion protein (66 kDa), induced by IPTG, was highly expressed in the form of inclusion body. G protein-specific polyclonal antibody was prepared using Japanese white rabbits, which were immunized with the purified recombinant G protein. The titer of the polyclonal antibody was up to 1:256000 detected by enzyme linked immunosorbent assay (ELISA). Western blotting and indirect immunofluorescent assay proved that the polyclonal antibody had high specificity, and neutralization test of polyclonal antibody showed that it could effectively prevent the cell from virus infection. This study lays the foundation for serological diagnosis method of SVCV and the research of SVCV pathogenic mechanism. |
| 罗培骁,张琪,王敏,刘学芹.2014.鲤春病毒血症病毒G蛋白的原核表达及多克隆抗体的制备[J].水生态学杂志,35(4):81-86. |
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