SFTSV糖蛋白Gn重组质粒的构建及其体液免疫原性研究  

Construction of recombinant plasmids expressing SFTSV glycoprotein Gn and immunogenicity study

作　　者：徐菱遥 韩亚萍[1] 周宜庆 金柯[1] 艾宇洁 黄祖瑚[1] 李军[1] 

机构地区：[1]南京医科大学第一附属医院感染病科,江苏南京210029

出　　处：《南京医科大学学报（自然科学版）》2016年第5期554-558,共5页Journal of Nanjing Medical University(Natural Sciences)

基　　金：国家重大科技专项(卫生部十二五支撑计划)(2013ZX10002005)

摘　　要：目的 :研究严重发热伴血小板减少综合征病毒(severe fever with thrombocytopenia syndrome virus,SFTSV)糖蛋白Gn的体液免疫原性。方法:将PCR方法扩增的SFTSV糖蛋白Gn基因和密码子优化后Gn基因克隆入载体p JW4303,构建Gn野生型和双优化重组质粒;经酶切和测序鉴定确认为目的质粒后,分别转染HEK293T细胞,Western blot检测糖蛋白Gn的表达;用Gn重组表达质粒及空载体分别免疫BALB/c小鼠,ELISA检测免疫后小鼠血清抗Gn特异性Ig G抗体。结果:成功构建Gn重组野生型质粒p JW4303-WSP-Gn和双优化质粒p JW4303-t PA-Gn-opt;Western blot证实p JW4303-WSP-Gn编码Gn可在HEK293T细胞内表达,p JW4303-t PA-Gn-opt编码Gn在HEK293T细胞内表达并分泌到细胞外;ELISA检测免疫后小鼠血清抗Gn特异性Ig G抗体证实各重组质粒均能诱导特异性Ig G抗体产生,双优化质粒较野生型质粒诱导产生的Ig G时间更早、滴度更高。结论:SFTSV的糖蛋白Gn具有良好的免疫原性;和野生型质粒相比,双优化质粒更有利于糖蛋白Gn的表达和分泌,并且具有更好的体液免疫原性。Objective：To explore the immunogenicity of severe fever with thrombocyte-penia syndrome virus（SFTSV）glycoprotein Gn. Methods：SFTSV glycoprotein Gn gene and optimal Gn gene were amplified by polymerase chain reaction（PCR）respectively. The gene product were cloned to p JW4303 to construct recombinant plasmids and all constructs were identified by sequencing,and then transiently transformed into HEK293 T cells to measure the Gn expression by Western blot. We immuned the BALB / c mice with recombinant plasmids and blank vector,and verified its immunogenicity by enzyme-linked immunosorbent method. Results：The recombinant plasmids p JW4303-WSP-Gn and p JW4303-t PA-Gn-opt were constructed success-fully. The p JW4303-WSP-Gn transiently expressed Gn antigen in cell lysates of HEK293 T cells in vitro,and p JW4303-t PA-Gn-opt in supernatants as same as cell lysates. The specific Ig G antibodies induced by p JW4303-t PA-Gn-opt was earlier and higher than that of p JW4303-WSP-Gn. Conclusion：SFTSV glycoprotein Gn showed good immunogenicity. The recombinant plasmid p JW4303-t PA-Gn-opt promoted the synthesis and secretion of Gn,and showed better humoral immunogenicity than p JW4303-WSP-Gn.

关 键 词：SFTSV 糖蛋白Gn 重组质粒 体液免疫原性 

分 类 号：R511[医药卫生—内科学]