戊型肝炎病毒抗原基因片段及其组合表达质粒的构建  

Construction of Plasmids Expressing Hepatitis E Virus Antigen Fragments and Their Combinations

作　　者：唐浩[1] 马雁冰[1] 李春宏[1] 杜瑞娟[1] 乐耕耘[1] 庄俊英[1] 刘勇[1] 孙茂盛[1] 戴长柏[1] 

机构地区：[1]中国医学科学院中国协和医科大学医学生物学研究所分子生物室,昆明650118

出　　处：《中国医学科学院学报》2000年第6期562-565,共4页Acta Academiae Medicinae Sinicae

摘　　要：目的选择及组合戊肝病毒 (HEV)抗原基因片段 ,进行表达与免疫学特性研究。方法根据已公布的 HEV序列 ,选择三段优势抗原表位 ,经 RT- PCR扩增基因片段 ,产物以单独或经甘氨酸连接肽串联组合的方式克隆到载体 pThioHis C中 ,经 DNA序列分析后 ,将构建的重组质粒转入大肠杆菌中进行表达 ,并以 Western印迹分析初步考察其免疫学特性。结果构建的六个重组质粒在大肠杆菌中获得了不同程度的表达 ,表达的目的蛋白具有与戊型肝炎患者阳性血清特异性结合的能力。结论 HEV抗原片段及其组合在大肠杆菌中得到了高效表达 ,并为优良诊断试剂及基因工程疫苗的研究提供了基础。Several different regions of the HEV antigen were expressed in Escherichia coli and their immunological characteristics were simply evaluated. Methods Three coding sequences of immunodominant antigenic regions in structural protein of the hepatitis E virus (ORF2.1: 6287- 6403nt, ORF2.2: 6743- 7126nt, ORF3: full length of ORF3) were amplified by RT- PCR from the fecal supernatants of macaques experimentally infected with HEV. These three fragments were inserted into the pThioHisC expression vector seperately or fused in line by (Gly)n short arms. After identified by DNA sequence analysis, these six recombinant plasmids were transformed into Escherichia coli, and immunology characteristics of expressed antigen fragments were evaluated by Western blotting. Results All six genes were successfully expressed in Escherichia coli. Western blotting assay showed that the recombinant proteins specifically reacted with the serum antibody from hepatitis E patient in various degrees. Conclusion Artificial antigen based on stringed epitops provides a promising strategy for detection and prevention of HEV infection.

关 键 词：基因克隆 大肠杆菌 基因表达 戊型肝炎病毒 

分 类 号：R373.21[医药卫生—病原生物学]