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目的 Mamu-B*007:03-sv1是中国恒河猴主要组织相容性复合体(major histocompatibility complex,MHC)I类分子的一种剪切异构体,其α3结构域缺失。本实验初步获得其在细胞内的表达和定位信息,并与全长的Mamu-B*007:03基因的表达情况进行比较。方法分别构建Mamu-B*007:03-sv1-myc-pEGFPN3和Mamu-B*007:03-myc-pEGFPN3的真核表达载体,并将这两种重组质粒分别转染293T细胞。利用Western-blot免疫印迹实验检测这两种基因在细胞内的表达情况,并利用激光共聚焦显微镜技术分别分析这两种基因在细胞内的表达定位。结果 Mamu-B*007:03-sv1和Mamu-B*007:03基因均能在293T细胞内正常表达。Mamu-B*007:03-sv1发生了糖基化的修饰,Mamu-B*007:03基因在细胞膜上表达,Mamu-B*007:03-sv1基因在细胞内表达。结论 Mamu-B*007:03-sv1基因的α3结构域缺失,其细胞内的表达和定位与全长Mamu-B*007:03基因有明显差异,其功能有待于进一步探索。
OBJECTIVE: Mamu-B * 007: 03-sv1 is a cleavage isoform of the major histocompatibility complex (MHC) class I in Chinese rhesus macaques with a3 domain deletion. In this experiment, the expression and localization information in the cells was preliminarily obtained and compared with the expression of the full-length Mamu-B * 007: 03 gene. Methods The eukaryotic expression vectors Mamu-B * 007: 03-sv1-myc-pEGFPN3 and Mamu-B * 007: 03-myc-pEGFPN3 were constructed and transfected into 293T cells respectively. Western-blot Western blotting was used to detect the expression of these two genes in the cells, and the expression localization of the two genes in the cells was analyzed by laser scanning confocal microscopy. Results Both Mamu-B * 007: 03-sv1 and Mamu-B * 007: 03 genes could be expressed normally in 293T cells. Glycosylation modification of Mamu-B * 007: 03-sv1 occurred. The Mamu-B * 007: 03 gene was expressed on the cell membrane and the Mamu-B * 007: 03-sv1 gene was expressed in the cell. Conclusion The deletion of the α3 domain of Mamu-B * 007: 03-sv1 gene and the expression and localization of the Mamu-B * 007: 03 gene in E. coli have a significant difference. The function of Mamu-B * 007: 03-sv1 gene remains to be further explored.