| 细粒棘球蚴抗原EPC1基因的克隆、表达及其免疫诊断的研究 |
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| 作者姓名： | Cai HX Shen YJ Han XM Yuan ZY Wang H Xu YX Hu Y Lu WY Guan YY Cao JP |
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| 作者单位： | 中国疾病预防控制中心寄生虫病预防控制所卫生部寄生虫病原与媒介生物学重点实验室世界卫生组织疟疾、血吸虫病和丝虫病合作中心;青海省地方病预防控制所; |
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| 基金项目： | 国家传染病科技重大专项(No.2009ZX10004-201,2008ZX10004-002)~~ |
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| 摘 要： | 目的克隆、表达细粒棘球蚴EPC1(EgEPC1)基因,鉴定重组抗原的反应原性,并用棘球蚴病患者血清评价其诊断价值。方法从绵羊肝脏棘球蚴囊中分离原头节,提取其总RNA,采用RT-PCR扩增EgEPC1基因,将其克隆至pGEM-T载体,再将其与原核表达载体PET28a(+)连接构建重组质粒PET28a-EgEPC1,转化大肠埃希菌(E.coli)BL21(DE3),经异丙基-β-D-硫代半乳糖苷(IPTG)诱导表达重组蛋白,用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)和蛋白质印迹(Western blotting)对纯化的重组抗原进行鉴定分析。以该重组蛋白作为包被抗原建立ELISA方法,检测细粒棘球蚴病患者血清(60份)特异性IgG抗体,同时以多房棘球蚴病(37份)、囊尾蚴病(16份)、华支睾吸虫病(7份)、日本血吸虫病(4份)患者和健康人血清(33份)作为对照,评价重组抗原EgEPC1的免疫诊断效果。结果双酶切鉴定和测序结果均显示重组质粒PET28a-EgEPC1构建成功。SDS-PAGE和Western blotting分析显示,重组质粒PET28a-EgEPC1在E.coli BL21(DE3)...
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| 关 键 词： | 细粒棘球绦虫 EPC1基因 基因表达 免疫诊断 |
Cloning, expression and immunodiagnostic evaluation of antigen EPC1 from Echinococcus granulosus |
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| Cai HX,Shen YJ,Han XM,Yuan ZY,Wang H,Xu YX,Hu Y,Lu WY,Guan YY,Cao JP.Cloning, expression and immunodiagnostic evaluation of antigen EPC1 from Echinococcus granulosus[J].Chinese Journal of Parasitology and Parasitic Diseases,2011,29(3):167-171. |
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| Authors: | Cai Hui-Xia Shen Yu-Juan Han Xiu-Min Yuan Zhong-Ying Wang Hu Xu Yu-Xin Hu Yuan Lu Wei-Yuan Guan Ya-Yi Cao Jian-Ping |
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| Institution: | CAI Hui-xia1,2,SHEN Yu-juan1,HAN Xiu-min2,YUAN Zhong-ying1,WANG Hu2,XU Yu-xin1,HU Yuan1,LU Wei-yuan1,GUAN Ya-yi1,CAO Jian-ping1 *(1 National Institute of Parasitic Diseases,Chinese Center for Disease Control and Prevention,Key Laboratory of Parasite and Vector Biology,MOH,WHO Collaborating Center of Malaria,Schistosomiasis and Filariasis,Shanghai 200025,China,2 Qinghai Institute for Endemic Disease Prevention and Control,Xining 811602,China) |
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| Abstract: | Objective To clone and express EPC1 gene of Echinococcus granulosus,and investigate its immun-ogenicity and diagnostic value.Methods Total RNA was extracted from hydatid cyst protoscoleces and EPC1 gene of Echinococcus granulosus was amplified by RT-PCR.The PCR product was cloned into pGEM-T vector,and then subclo-ned into the prokaryotic expression vector PET28a(+).The positive recombinants were transformed into Escherichia coli BL21(DE3),and followed by expression of the protein induced by IPTG.The recomb... |
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| Keywords: | Echinococcus granulosus EPC1 gene Gene expression Immunodiagnosis |
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