牛丙型肝炎病毒核心蛋白结合蛋白6同源基因的克隆化研究

目的　利用不同种属动物之间重要基因序列高度同源的理论 ,应用生物信息学技术和方法 ,克隆牛丙型肝炎病毒 (HCV)核心蛋白结合蛋白 6 (HCBP6 )的同源基因。方法　首先应用酵母双杂交技术 ,以表达HCV核心蛋白的表达载体为诱饵 ,对于百万级的肝细胞cDNA文库酵母进行配合、筛选 ,首先获得人HCBP6的全长编码基因 ,然后应用美国国立生物信息学中心 (NCBI)建立的核苷酸序列数据库 (GenBank)的同源基因的检索 ,搜索与之同源的来源于牛的表达序列标签 (EST)。然后根据基因同源性的原则 ,确定牛HCBP6的同源基因。结果　获得了与HCV核心蛋白结合蛋白的 36个基因片段 ,其中之一命名为HCBP6。根据基因同源性搜索 ,获得了来源于牛的EST基因序列片段 ,最终确立了牛HCBP6的同源基因。结论　利用不同物种之间基因同源性的原理 ,NCBI数据库GenBank同源基因的搜索 ,获得了牛HCBP6同源基因。生物信息学技术在后基因组时代具有重要地位和作用。

Cloning and sequence analysis of Bos taurus hepatitis C virus core-binding protein 6 (HCBP6) homologue gene

Aim To clone Bos taurus hepatitis C virus (HCV) core protein-binding protein 6 (HCBP6)homologue gene by homologous gene searching in the GenBank established by National Center of Bioinformatics Institute (NCBI),National Institute of Health(NIH),USA.Methods The expressive vector of HCV core protein was constructed.The transformed yeast was mated with yeast harboring hepatocyte cDNA library.The positive clones were selected by galactosidase activities and deficient medium growth.The homologous expressed sequence tag (EST) were searched in the GenBank for Bos taurus cDNA fragments.The full length coding sequence for Bos taurus HCBP6 homologue was established by bioinformatics method.Results Thirty-six positive clones have been selected from yeast-two hybrid screening.Among them,there were 3 cDNA fragment without homology to any sequences in the GenBank.Huamn HCBP6 cDNA has been cloned.The Bos taurus HCBP6 full length coding sequence has been finished by homologous searching.Conclusion The Bos taurus HCBP6 full length coding sequence haw been established by combination of molecular cloning and bioinformatics methods.Bioinformatics method is an important tool for identification and characterization of homologous genes from GenBank in the post-genomic era.