Optimization of heteroduplex analysis for the detection of BRCA mutations and SNPs

Abstract

BRCA1 and BRCA2 are tumour suppressor genes whose mutant phenotypes predispose to breast and
ovarian cancer. Screening for mutations in these genes is now standard practice for hereditary breast and ovarian cancer
(HBOC) cases in Europe, and permits medical follow-up and genetic counselling adapted to the needs of individuals in
such families. Currently, most laboratories performing diagnostic analysis of the BRCA genes use PCR of exons and
intron-exon boundaries coupled to a pre-screening step to identify anomalous amplicons. The techniques employed for
the detection of mutations and SNPs have evolved over time and vary in sensitivity, specificity and cost-effectiveness. As
a variant for pre-screening techniques, we chose the recently developed Surveyor® heteroduplex cleavage method as a
sensitive and specific technique to reveal anomalous amplicons of the BRCA genes, using only basic laboratory
equipment and agarose gel electrophoresis. Here we present the detection of either mutations or SNPs within the BRCA1
exon 7, using heteroduplex analysis (HA) by mismatch-specific endonuclease, confirmed by dideoxy sequencing.