猪细胞巨化病毒四川株MCP基因克隆及其编码蛋白的生物信息学分析  

Cloning MCP gene of porcine cytomegalovirus Sichuan strain and bioinformatics analysis of its coding protein

作　　者：史小红[1,2] 徐志文[1,2] 郭万柱[1,2] 朱玲[1,2] 漆信桥[1,2] 王燕群[1,2] 赵玲[1,2] 

机构地区：[1]四川农业大学动物生物技术中心,四川雅安625014 [2]四川农业大学动物疫病与人类健康实验室,四川雅安625014

出　　处：《中国兽医学报》2011年第11期1559-1563,共5页Chinese Journal of Veterinary Science

基　　金：国家十一五科技支撑计划资助项目(2006BAD06A18);"长江学者和创新团队发展计划"创新团队项目(IRT0848);四川省青年科技基金项目(2006Q14-049)

摘　　要：参照GenBank公布的仅有的日本株猪细胞巨化病毒较大衣壳蛋白(MCP)基因序列(登录号:AB051069)设计2对引物,采用分步克隆的方法,将PCMV MCP全序列克隆入pMD19-T载体进行测序,成功获得了pMD-MCP重组质粒,将所得序列录入到GenBank中(登录号:HQ025802)。测序结果表明,该MCP基因全长4 017bp,共编码1 338个氨基酸;与NCBI上公布的仅有的日本毒株MCP基因的核苷酸同源性为96.8%,氨基酸同源性为94.1%;进化分析显示:PCMV MCP基因与人疱疹病毒6型或7型的MCP基因亲缘关系较近;利用生物信息学软件对蛋白质结构特征进行分析,发现该蛋白含有59个潜在的磷酸化位点,潜在功能强大;亚细胞定位预测结果表明该蛋白主要存在于线粒体中和细胞质中并各占39.1%和26.1%,内质网占17.4%,高尔基体占8.7%,空泡和细胞核均占4.3%,表明PCMV MCP蛋白属于胞质蛋白,抗原区位集中于胞膜,有向胞质移动的趋势;另该成熟蛋白存在18个主要的抗原位点,将肽链经亲水性与抗原表位的共同分析,发现其肽链的C端极有可能分布有抗原决定簇。According to the only genomic sequence of PCMV MCP gene from Japan （GenBank accession number： AB051069）, two pairs of specific primer were designed to amplify the fragment encoding PCMV MCP gene by PCR. These fragment was cloned into pMD19-T vector to sequence. The recombinant plasmid was designated as pMD- MCP successfully. The results indicated that length of PCR product of interest was 4 017 bp,encoding 1 338 amino acids in all. the alignment result showed that the homology of nucleotide was 96.8% and the homology of amino acid sequences was 94.1 ~/00 with the published sequence of Japan strains from GenBank. In phylogenetic tree analysis with the standard genotype sequence of GenBank,the genome was clustered with HHV-6 and HHV-7. Using the bioinfor- matics software and database to analyze the PCMV MCP gene and its encoding protein,the results suggested that it involved 59 potential phosphorylation sites, indicating that this peptide chain has unlimited potential. The mature protein located mainly in mitochondriaI, cytoplasmic, and endoplasmic reticulum accounting for 39. 1%, 26. 1 % and 17.4% ,respectively,minor in Golgi,vacuolar and nuclear for 8.7 % and 4. 3%, respectively. It was conformed that the encoded protein,which was cytoplasmic protein and its antigenic sites was on the surface of membrane to shift slowly to cytoplasmic. Otherwise the mature protein included 18 principal antigenic sites, according to the result of hydrophilic prediction the antigenic determinant in the C-terminal of the peptide chain was fond. The full MCP gene coding region of PCMV virus isolated in China （GenBank accession number： HQ025802） was sequenced first time. It could be used for study on the package,mature,stability and immune process of the virus.

关 键 词：猪细胞巨化病毒 主要衣壳蛋白基因 克隆 生物信息学分析 

分 类 号：S852.65[农业科学—基础兽医学]