Identification by a random linker insertion method of a region which suppresses the transforming activity of the raf oncogene |
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Authors: | F Ishikawa R Sakai M Ochiai T Sugimura M Nagao |
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Institution: | Carcinogenesis Division, National Cancer Center Research Institute, Tokyo, Japan. |
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Abstract: | Activation mechanism of raf oncogene was studied by applying in vitro mutagenesis to its cDNA. Previous studies suggested the presence of an activation suppressing sequence in the amino-terminal half of c-raf product. Loss of the sequence by genetic rearrangement was presumed to convert c-raf to possess transforming activity. To identify such sequence, we prepared cDNA mutants by random linker insertion. Synthetic oligonucleotide linker was inserted into the plasmid containing cDNA at a single and random site. Coupling two different mutants, in-frame deletion mutants were constructed systematically. Analysis of these deletion mutants revealed a region, the loss of which made c-raf activated. |
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