北京地区两株人偏肺病毒M蛋白基因的原核表达及抗原活性分析  

Prokaryotic Expression and Antigenic Activity Analysis on the Matrix Protein Genes of Two Strains of Human Metapneumovirus Recently Identified in Beijing

作　　者：曹守春[1] 钱渊[1] 李国华[1] 朱汝南[1] 赵林清[1] 丁雅馨[1] 

机构地区：[1]首都儿科研究所病毒研究室北京市感染与免疫中心实验室,北京100020

出　　处：《病毒学报》2007年第1期60-62,共3页Chinese Journal of Virology

基　　金：国家自然科学基金(30570080);北京市自然科学基金(7052020)

摘　　要：Human metapneumovirus(hMPV) is a recently identified respiratory virus more like human respiratory syncytial virus in clinical symptoms.Matrix protein(M) is one of the most important structural proteins.For further studying of hMPV,the full length of M genes from the recombinant plasmid pUCm-M1816 and pUCm-M1817 were cloned by PCR and sub-cloned into the pET30a(+) vector,which is a prokaryotic expression vector,after dual-enzyme digestion with Bam HI and Xho I.The positive recombinated plasmids were transformed into E.coli BL21(DE3) and expressed under the inducing of IPTG.Target proteins were characterized by SDSPAGE and Western blotting.In this article,we’ve successfully constructed the recombinated plasmids pET30a-M1816 and pET30a-M1817 which have correct open reading frames confirmed by dual-enzyme digestion analysis and sequencing. The fusion proteins with 6·His-N were highly produced after inducing by 1mmol/L IPTG at 37℃.A unique protein band with approximate 27.6 kD was characterized by SDS-PAGE.Most of the target protein existed in inclusion body.Western blot analysis showed that the target protein has specific binding reaction to rabbit antiserum against polypeptides of the matrix protein of hMPV.So the M genes were highly expressed in the prokaryotic system and the expressed M proteins have specific antigenic activities.It can be used for further studying of hMPV infections in Beijing.Human metapneumovirus （hMPV） is a recently identified respiratory virus more like human respiratory syncytial virus in clinical symptoms. Matrix protein （M） is one of the most important structural proteins. For further studying of hMPV, the full length of M genes from the recombinant plasmid pUCm-M1816 and pUCm- M1817 were cloned by PCR and sub-cloned into the pET30a（ ＋ ） vector, which is a prokaryotic expression vector, after dual-enzyme digestion with Barn HI and Xho Ⅰ. The positive recombinated plasmids were transformed into E. coli BL21 （DE3） and expressed under the inducing of IPTG. Target proteins were characterized by SDS-PAGE and Western blotting. In this article, we＇ ve successfully constructed the recombinated plasmids pET30a-M1816 and pET30a-M1817 which have correct open reading frames confirmed by dual-enzyme digestion analysis and sequencing. The fusion proteins with 6·His-N were highly produced after inducing by 1 mmol/ L IPTG at 37℃. A unique protein band with approximate 27.6 kD was characterized by SDS-PAGE. Most of the target protein existed in inclusion body. Western blot analysis showed that the target protein has specific binding reaction to rabbit antiserum against polypeptides of the matrix protein of hMPV. So the M genes were highly expressed in the prokaryotic system and the expressed M proteins have specific antigenic activities. It can be used for further studying of hMPV infections in Beijing.

关 键 词：人偏肺病毒 基质蛋白M 原核表达 抗原活性分析 

分 类 号：R373.1[医药卫生—病原生物学]