Hierarchical mutation screening protocol for the BRCA1 gene

The identification of mutations in the BRCA1 gene poses difficulties in achieving a screening outcome that satisfies the twin needs of speed and accuracy. These needs must also take into account the patient's family history and the statistical evaluation of the probability of detecting a mutation. Given the above, we present here a hierarchical mutation screening strategy that comprises two tiers: first, multiplex heteroduplex and exon 13 duplication analysis; second, exon amplification and direct sequencing using a 96-well tray format. The advantages of this strategy are two-fold: first, the division of analytical tools in order to achieve low and high-resolution mutation screening, respectively; second, a streamlined sequencing approach that leads to a sensitive and rapid assay that reduces labor costs and handling errors. The success of this approach is shown by the identification of a novel deletion mutation in exon 14 of the BRCA1 gene, which was not detected by the more conventional protein truncation assay due to the small size of the predicted truncated protein.