Fujita J, Yoshida O, Ebi Y, Nakayama H, Onoue H, Rhim J S, Kitamura Y
Institute for Cancer Research, Osaka University Medical School, Japan.
Urol Res. 1988;16(6):415-8. doi: 10.1007/BF00280021.
To detect mutationally activated ras oncogenes, we analyzed electrophoretic mobilities of ras p21 proteins utilizing the fact that many ras oncogenes produce abnormal p21 proteins that migrate at SDS/polyacrylamide gel electrophoresis as a fast-moving or slow-moving species in comparison to a normal p21 depending on the kind of mutation. Of 18 human tumor cell lines analyzed, four (SW480, SW620 and SW403 colon cancers, and SW626 ovary cancer) produced p21 belonging to the slow-moving species, suggesting a point mutation within codon 12 of a member of the three ras genes, H-, Ki- and N-ras. Subsequent DNA transfection analysis using NIH/3T3 cells as recipients identified activated Ki-ras oncogenes in the same four but not in other 14 cell lines. Thus, the analysis of p21 might serve as a rapid primary method to screen a large number of tumor materials for the presence of certain types of mutationally activated ras oncogenes.
为了检测发生突变激活的ras癌基因，我们利用许多ras癌基因会产生异常p21蛋白这一事实，分析了ras p21蛋白的电泳迁移率。与正常p21相比，这些异常p21蛋白在SDS/聚丙烯酰胺凝胶电泳中表现为快速迁移或慢速迁移的条带，具体取决于突变的类型。在分析的18个人类肿瘤细胞系中，有四个（SW480、SW620和SW403结肠癌以及SW626卵巢癌）产生属于慢速迁移条带的p21，这表明三个ras基因（H-、Ki-和N-ras）之一的第12密码子内存在点突变。随后以NIH/3T3细胞作为受体进行的DNA转染分析在同样的四个细胞系中鉴定出激活的Ki-ras癌基因，而在其他14个细胞系中未鉴定出。因此，对p21的分析可作为一种快速的初步方法，用于筛查大量肿瘤材料中是否存在某些类型的发生突变激活的ras癌基因。
