Inverse restriction site mutation (iRSM) analysis. Mutation detection involving the formation of restriction enzyme sites in target genes

This paper describes a rapid screening procedure for the detection of DNA sequence changes resulting in the creation of new restriction enzyme sites. The basic methodology involves the identification of the conversion of one restriction site into another by mutagenesis. The selective removal of the wild-type sequences by digestion with a restriction enzyme acting on the wild-type sequence increases the sensitivity beyond that of PCR-RFLP analysis (10(-4)-10(-5) detectable here). In this paper we describe the rapid detection of induced in vivo mutations transforming the ApaI restriction site present in intron 6 of the mouse p53 gene to a unique AvaII site. The potential application of this method in other genes and organisms as a rapid screen for induced mutations is discussed.