H5亚型禽流感病毒HA基因的克隆、表达及其生物学活性的初步分析  

Expression of Hemagglutinin Gene of Avian influenza virus Subtype H5 in Escherichia coli and the Preliminary Analysis of Bioactivity

作　　者：武山[1] 陈晨[1] 雷霆[1] 曹红[1] 陈福勇[1] 

机构地区：[1]中国农业大学动物医学院,北京100094

出　　处：《农业生物技术学报》2005年第6期739-742,共4页Journal of Agricultural Biotechnology

摘　　要：参考GenBank上发表的H5亚型禽流感病毒(Avianinfluenzavirus,AIV)血凝素(HA)基因序列设计引物,经RT-PCR扩增HA基因的HA1片段,再将其克隆到原核表达载体PET-28a中,用大肠杆菌(Escherichiacoli)BL21(DE3)原核表达,SDS-PAGE和Westernblot对表达蛋白进行鉴定。Westernblot结果表明,表达蛋白能与H5亚型AIV单特异性血清特异性反应。交叉反应试验表明,此蛋白具有良好的抗原性,可被H5亚型AIV抗体特异性识别且无交叉反应。用变性纯化后的蛋白建立了对AIV抗体检测的间接ELISA,并对236份鸡血清样本进行了实验室检测。检测结果与血凝抑制试验检测结果符合率达到100%。综合研究表明,表达蛋白具有良好抗原性。The hemagglutinin（HA） HA1 gene of Avian intluenza virus （AIV） subtype H5 was amplified by RT-PCR according to the sequence published on GenBank. It was cloned into the expression vector PET-28a. The protein encoded by this gene was expressed in Escherichia coli strain BL21 （DE3） and identified by SDS-PAGE and Westem-blot. The results indicated that this protein could be specifically recognized by H5 subtype AIV antiserum. The cross-reactivity test results showed that this protein could only react with H5 subtype AIV antiserum but not with any other antiserum against NDV, IBV, etc. The results from detection of 236 serum samples by ELISA using the purified protein were 100% in agreement with that of HI test. All these results demonstrate that this expressed protein is of good antigenicity and can hold great promise for development of diagnostic kits for avian influenza.

关 键 词：禽流感病毒 H5亚型 血凝素(HA) 原核表达 ELISA 

分 类 号：S858.305.3[农业科学—临床兽医学] S852.659.5[农业科学—兽医学]