DETECTION OF MUTATIONS IN GC-RICH DNA BY BISULFITE DENATURING GRADIENT GEL-ELECTROPHORESIS

Denaturing gradient gel electrophoresis (DGGE) in combination with PCR and 'GC-clamping' has proven highly efficient as a method for detecti on of DNA sequence differences. Due to strand dissociation phenomena, however, its use has been limited to the analysis of sequences with a relatively low content of GC pairs. This paper describes how treatment of template DNA with sodium bisulphite drastically lowers the melting temperature of very GC-rich sequences and renders them amenable to DG GE analysis. We demonstrate the use of bisulphite DGGE for rapid and e fficient detection of mutations in the p16(INK4/CDKN2) tumour suppress or gene.