Gilmer T M
Mol Cell Biol. 1984 May;4(5):846-51. doi: 10.1128/mcb.4.5.846-851.1984.
The cellular homolog of the Rous sarcoma virus transforming gene (v-src) was cloned into a plasmid containing the simian virus 40 origin of replication and transcriptional signals. This recombinant plasmid, designated pSVOHCS11 , directs the synthesis of relatively high levels of c-src mRNA and c-src protein ( pp60c -src), when the plasmid is studied 48 to 72 h after calcium phosphate-mediated DNA transfection of COS (monkey) cells. The level of c-src mRNA synthesis is 50-fold higher than the amount of c-src RNA produced in uninfected chicken embryo fibroblasts. Furthermore, the level of pp60c -src expressed in pSVOHCS11 -transfected COS cells is approximately the same as that of pp60v -src in Rous sarcoma virus-transformed cells. Using this recombinant plasmid, we demonstrated that c-src mRNA contains sequences which map 3' to the previously identified c-src-v-src regions of homology. In view of the small amount of c-src mRNA and protein that can be isolated from uninfected cells, this transient expression system offers a convenient source of material for further analyses of the c-src gene product.
劳氏肉瘤病毒转化基因(v-src)的细胞同源物被克隆到一个含有猴病毒40复制起点和转录信号的质粒中。这个重组质粒命名为pSVOHCS11,当在磷酸钙介导的DNA转染COS(猴)细胞48至72小时后研究该质粒时,它指导合成相对高水平的c-src mRNA和c-src蛋白(pp60c-src)。c-src mRNA的合成水平比未感染的鸡胚成纤维细胞中产生的c-src RNA量高50倍。此外,在pSVOHCS11转染的COS细胞中表达的pp60c-src水平与劳氏肉瘤病毒转化细胞中的pp60v-src水平大致相同。利用这个重组质粒,我们证明c-src mRNA含有位于先前鉴定的c-src-v-src同源区域3'端的序列。鉴于从未感染细胞中可分离到的c-src mRNA和蛋白量很少,这个瞬时表达系统为进一步分析c-src基因产物提供了一个方便的材料来源。