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Mutagenesis, Vol. 14, No. 1, 37-42, January 1999
© 1999 UK Environmental Mutagen Society/Oxford University Press

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

Gareth J.S. Jenkins1, Nobuo Takahashi2 and James M. Parry

Centre for Molecular Genetics and Toxicology, University of Wales Swansea, Singleton Park, Swansea SA2 8PP, UK

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.

1 To whom correspondence should be addressed. Tel: +44 1792 205678. Fax: +44 1979 295447; Email: g.j.jenkins{at}swansea.ac.uk

2 Present address: Otsuka Pharmaceutical Co. Ltd, 463-10 Kagasuno Kawauchi-cho, Tokushima 771-01 Japan


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