Fragile-X syndrome is a hereditary dynamic mutation disorder, predominantly caused by a large expansion of CGG trinucleotide repeats in the FMR 1 gene leading to methylation and down regulation of transcription of the gene. For the molecular diagnosis of the disease, the repeat locus in FMR 1 gene is primarily detected by Southern blotting with radioactively labeled probes. Unusually GC rich composition of the expanded region caused technical difficulties during PCR based testing and therefore is not performed by most diagnostic laboratories. We established a Southern blot method and a novel PCR protocol which enables the amplification of normal, premutated and full mutated alleles. In both techniques bands are visualized by digoxigenin labeled probe and chemiluminescent detection. Although diagnosis of fragile-X was possible by cytogenetic analysis in our division, it was not possible to diagnose premutation carriers or mosaic fragile-X patients. This newly established nonradioactive PCR and Southern blot analysis will provide routine detection of full fragile-X mutations and premutations in our laboratory and therefore enable us to offer accurate genetic counseling and prenatal diagnosis.

Fragile-X syndrome is a hereditary dynamic mutation disorder, predominantly caused by a large expansion of CGG trinucleotide repeats in the FMR 1 gene leading to methylation and down regulation of transcription of the gene. For the molecular diagnosis of the disease, the repeat locus in FMR 1 gene is primarily detected by Southern blotting with radioactively labeled probes. Unusually GC rich composition of the expanded region caused technical difficulties during PCR based testing and therefore is not performed by most diagnostic laboratories. We established a Southern blot method and a novel PCR protocol which enables the amplification of normal, premutated and full mutated alleles. In both techniques bands are visualized by digoxigenin labeled probe and chemiluminescent detection. Although diagnosis of fragile-X was possible by cytogenetic analysis in our division, it was not possible to diagnose premutation carriers or mosaic fragile-X patients. This newly established nonradioactive PCR and Southern blot analysis will provide routine detection of full fragile-X mutations and premutations in our laboratory and therefore enable us to offer accurate genetic counseling and prenatal diagnosis.