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为了研究斜纹夜蛾核型多角体病毒Ⅱ型(Splt MNPVⅡ)中ORF117基因(编码GP117蛋白)的结构和功能,根据其核苷酸序列和氨基酸序列进行了生物信息学分析,以此为基础对该基因的启动子活性和转录时相进行分析,表达纯化了GP117蛋白的部分序列以制备豚鼠来源多克隆抗体,并利用该抗体对ORF117基因的表达时相和GP117蛋白在细胞中的定位进行了分析。生物信息学分析表明该基因全长1 203 bp,编码400个氨基酸的蛋白质,预测分子量为45.5 k Da,等电点为5.06;启动子活性和转录时相分析结果表明ORF117是一个晚期表达的基因;原核表达抗原免疫制备的多克隆抗体效价可以达到1∶3 200,利用该抗体对其表达时相的分析进一步验证了ORF117为晚期表达基因,免疫荧光检测表明编码的蛋白多存在于细胞质中。
In order to study the structure and function of ORF117 (encoding GP117 protein) in Splt MNPV Ⅱ, bioinformatics analysis based on its nucleotide and amino acid sequences was carried out. Based on this, The promoter activity and transcriptional phase of this gene were analyzed. The partial sequence of GP117 protein was expressed and purified to prepare the guinea pig-derived polyclonal antibody, and the expression of ORF117 gene and the localization of GP117 protein in the cell were performed by this antibody analysis. Bioinformatics analysis showed that the gene was 1 203 bp in length and encoded a protein of 400 amino acids with a predicted molecular weight of 45.5 k Da and an isoelectric point of 5.06. The promoter activity and transcriptional phase analysis indicated that ORF117 was a late-expressed gene . The titer of the polyclonal antibody immunoprecipitated with the prokaryotic expression antigen could reach 1: 3 200. The analysis of its expression phase by this antibody further confirmed that ORF117 is a late expression gene. Immunofluorescence showed that many of the encoded proteins existed in the cytoplasm .