Validation of a Screening Method for Dynamic Mutations in the Gene.

The objectives of this study were to validate the direct triplet-primed PCR method (dTP-PCR) for determination of dynamic mutations in the gene, and to compare the results of the dTP-PCR method and Southern blot analysis. The number of CGG repeats in the gene was determined by the direct triplet-primed PCR method and by melting curve analysis. The cut-off temperature between normal and permutations of the CGG repeats was determined using control samples with a known number of CGG repeats. All patients are classified into four categories based on the DNA melting curve. The clinical performance of the assay was established by 40 previously analyzed samples, yielding results of 100% sensitivity and 90.48% specificity in detection expansions of CGG (>30) repeats in the gene. This method is appropriate for the quick determination of allelic changes in the gene, screening a population, and identifying mutations or premutation carriers in a population with intellectual disabilities of an unknown cause.