| 高致病猪繁殖与呼吸综合征病毒JXA1株Nsp2基因分段克隆表达及其单克隆抗体的制备 |
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| 引用本文： | 胡鸿惠,曹振,赵铁柱,王斌,秦亚嫚,杨璐,王传彬,陈西钊,田克恭.高致病猪繁殖与呼吸综合征病毒JXA1株Nsp2基因分段克隆表达及其单克隆抗体的制备[J].中国畜牧兽医,2009,36(10):39-44. |
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| 作者姓名： | 胡鸿惠 曹振 赵铁柱 王斌 秦亚嫚 杨璐 王传彬 陈西钊 田克恭 |
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| 作者单位： | 胡鸿惠(中国农业大学动物医学院,北京,100193;中国动物疫病预防控制中心,北京,100125);曹振,赵铁柱,秦亚嫂,杨璐,王传彬,陈西钊,田克恭(中国动物疫病预防控制中心,北京,100125);王斌(北京市动物卫生监督所,北京,100044) |
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| 基金项目： | 国家科技支撑计划课题，国家863项目 |
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| 摘 要： | 分析猪繁殖与呼吸综合征病毒(porcine reproductive and respiratory syndrome virus,PRRSV)非结构蛋白Nsp2,扩增和克隆JXA1株Nsp2基因的4个片段.并将它们分别插人pGEx-6P-1表达载体构建重组表达质粒,经IPTG诱导表达得到蛋白,纯化后进行活性鉴定.以分段表达的蛋白为抗原,建立间接ELISA方法用于杂交瘤细胞株的筛选.用PRRSVNVDC-JXA1株灭活疫苗对BALB/c小鼠进行常规免疫,最后一次用活毒加强免疫,取其脾细胞与SP2/0细胞在聚乙二醇作用下融合,经过选择性培养、有限稀释筛选到1株针对Nsp2的杂交瘤细胞株688.鉴定结果表明,该株杂交瘤细胞仅与4个蛋白片段中的Nsp2F1有反应,细胞经长期体外培养和冻存后复苏能稳定分泌抗体,上清效价达1:210,腹水效价达1:106.本研究成功表达了4个PRRSV JXA1株的非结构蛋白Nsp2的片段,以其为抗原建立筛选方法,得到1株针对Nsp2的单克隆抗体,为研究Nsp2功能和建立相关诊断方法奠定物质基础.
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| 关 键 词： | 单克隆抗体 |
Expression and Development of Monoclonal Antibody against Nsp2 Gene Fragments of Highly Virulent PRRSV JXA1 Strain |
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| HU Hong-hui,CAO Zhen,ZHAO Tie-zhu,WANG Bin,QIN Ya-man,YANG Lu,WANG Chuan-bin,CHEN Xi-zhao,TIAN Ke-gong.Expression and Development of Monoclonal Antibody against Nsp2 Gene Fragments of Highly Virulent PRRSV JXA1 Strain[J].China Animal Husbandry & Veterinary Medicine,2009,36(10):39-44. |
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| Authors: | HU Hong-hui CAO Zhen ZHAO Tie-zhu WANG Bin QIN Ya-man YANG Lu WANG Chuan-bin CHEN Xi-zhao TIAN Ke-gong |
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| Institution: | HU Hong-hui1,2,CAO Zhen2,ZHAO Tie-zhu2,WANG Bin3,QIN Ya-man2,YANG Lu2,WANG Chuan-bin2,CHEN Xi-zhao2,TIAN Ke-gong2 (1.College of Veterinary Medicine,China Agricultural University,Beijing 100193,China,2.China Animal Disease Control Center,Beijing 100125,3.Beijing Animal Health Inspection Institute,Beijing 100044,China) |
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| Abstract: | Analyzing PRRSV gene Nsp2 by biological software,four gene fragments of Nsp2 were successfully amplified by RT-PCR and cloned from the PRRSV JXA1 strain.Their expressing plasmids were constructed by inserting the target gene fragments into pGEX-6P-1 vector and were induced with IPTG.Subsequently,the collecting and purifying proteins were identified.Using these proteins,we established an indirect ELISA for bolting McAbs.We immunized the BALB/c mice with PRRSV vaccine.After the final immunization with virus,s... |
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| Keywords: | PRRSV JXA1 Nsp2 |
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