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基于CMV启动子表达鹦鹉热衣原体PmpD-N蛋白的重组火鸡疱疹病毒的构建 预览
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作者 刘杉杉 孙伟 +5 位作者 张稳 贾长青 罗杰 沈易华 郁建生 何诚 《动物医学进展》 北大核心 2018年第7期13-18,共6页
为构建表达鹦鹉热衣原体多型外膜蛋白D的N端(PmpD-N)蛋白重组火鸡疱疹病毒(HVT),以鹦鹉热衣原体CB7株DNA为模板,通过PCR扩增pmpD-N基因,构建带有巨细胞病毒(CMV)启动子的真核表达盒,采用细菌人工染色体技术构建HVT重组子,转染细胞获得HV... 为构建表达鹦鹉热衣原体多型外膜蛋白D的N端(PmpD-N)蛋白重组火鸡疱疹病毒(HVT),以鹦鹉热衣原体CB7株DNA为模板,通过PCR扩增pmpD-N基因,构建带有巨细胞病毒(CMV)启动子的真核表达盒,采用细菌人工染色体技术构建HVT重组子,转染细胞获得HVT重组病毒,并在体外评价该重组病毒的相关特性。结果表明,转染细胞后成功获得HVT重组病毒。通过Westernblot和IFA分析表明,重组病毒表达了PmpD-N蛋白。Confocal分析表明,PmpD-N蛋白分布在细胞的胞核、胞浆和细胞表面。HVT重组病毒的复制特性和野生型HVT一致。HVT重组病毒能够激活巨噬细胞促炎性细胞因子IL-6和NO的产生。该试验成功构建了含有CMV启动子,能够表达PmpD-N蛋白的HVT重组病毒,为下一步进行动物试验和分析该病毒的免疫效力奠定了基础。 展开更多
关键词 鹦鹉热衣原体 多型外膜蛋白D的N 火鸡疱疹病毒 细菌人工染色体技术 巨细胞病毒启动子
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Development of a Novel PmpD-N ELISA for Chlamydia psittaci Infection 预览
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作者 LIU Shan Shan CHU Jun +3 位作者 ZHANG Qiang SUN Wei ZHANG Tian Yuan HE Cheng 《生物医学与环境科学:英文版》 SCIE CAS CSCD 2016年第5期315-322,共8页
Objective Chlamydia psittaci is an avian respiratory pathogen and zoonotic agent. The wide prevalence of C. psittaci poses a threat to the poultry industry and its employees. However, few commercial kits are available... Objective Chlamydia psittaci is an avian respiratory pathogen and zoonotic agent. The wide prevalence of C. psittaci poses a threat to the poultry industry and its employees. However, few commercial kits are available for detecting avian antibodies excluding the in-house ELISA kit. In this study, we developed a novel ELISA kit for detecting antibodies against C. psittaci based on the N-terminal fragment of polymorphic outer membrane protein D(Pmp D-N) as the coating antigen. Methods The antigen concentrations, primary antibody, and cut-off value were determined and optimized. The ELISA, designated Pmp D-N ELISA, was assessed for sensitivity, specificity, and concordance using sera samples from 48 experimentally infected and 168 uninfected SPF chickens. Results The sensitivity and specificity of Pmp D-N ELISA were 97.9%, 100%, respectively, while the concordance was 98.1% as compared to that of MOMP-ELISA. No cross-reaction with positive sera for other avian pathogens was found. Using Pmp D-N ELISA, 799/836 clinical samples were positive, including 93.0% and 98.1% positivity in layers and broilers, respectively. Conclusion These data indicate that indirect ELISA with Pmp D-N as the antigen candidate is a promising approach for the surveillance of C. psittaci infection. 展开更多
关键词 ELISA方法 鹦鹉热衣原体 衣原体感染 酶联免疫试剂盒 ELISA试剂盒 禽流感抗体 间接ELISA 阳性血清
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