Denaturing high-performance liquid chromatography detects reliably BRCA1 and BRCA2 mutations

Denaturing high-performance liquid chromatography (DHPLC) is a recently dev eloped method of comparative sequencing based upon heteroduplex detection. To assess the reliability of this method, 180 different mutations (54 delet ions, 12 insertions, and 117 single base substitutions) in BRCA1 and BRCA2 were tested. Second, 25 index individuals with complete DHPLC analysis of B RCA1 were reanalyzed by dye-terminator sequencing. Third, 41 index individu als were analyzed concomitantly by both DGGE and DHPLC, Of the 180 differen t BRCA1 and BRCA2 mutations, 179 showed heterozygous DHPLC elution profiles . Dye-terminator sequencing of the entire BRCA1 gene, including 5592 bp of coding sequence and 5206 bp of flanking noncoding sequence, in 25 index ind ividuals did not reveal additional variants missed by DHPLC, The concomitan t analysis of 41 index cases showed that 4 probably disease-associated muta tions were identified by DHPLC while only 3 of those 4 sites were detected by denaturing gradient gel electrophoresis. We conclude that DHPLC is a sen sitive and cost-effective method for the screening of BRCA1 and BRCA2. (C) 1999 Academic Press.