NONISOTOPIC ANALYSIS OF SINGLE-STRAND CONFORMATION POLYMORPHISM (SSCP) IN THE EXON-13 REGION OF THE HUMAN DYSTROPHIN GENE

More than 30% of Duchenne and Becker muscular dystrophy (DMD/BMD) pati ents. have no gross DNA rearrangements like deletions or duplications. The large size of the coding sequence of the dystrophin gene (11 kilo bases) complicates systematic identification of point mutations. Recen tly reported approaches based on genomic DNA or mRNA show that chemica l cleavage of mismatches is an effective but time consuming and techni cally demanding method for the identification of point mutations in th e human dystrophin gene. We have used a fast and convenient system con sisting of PCR amplification of genomic DNA, non-isotopic SSCP analysi s, and direct sequencing of PCR products for the detection of mutation s in exon 13 and adjacent intron sequences. Sixty-eight DMD patients w ithout detectable deletions or duplications were analysed, resulting i n the identification of a point mutation in the coding sequence and tw o polymorphisms in the 5' flanking intron. The C to T change of the fi rst nucleotide in the third triplet leads to a stop codon and seems to be the cause of the functional deficiency of the gene product in this patient.