CONSTANT DENATURANT GEL-ELECTROPHORESIS (CDGE) IN BRCA1 MUTATION SCREENING

Screening for mutations in the breast and ovarian cancer susceptibilit y gene, BRCA1,is complicated by the wide spectrum of mutations found i n this large gene, In the present study a constant denaturant gel elec trophoresis (CDGE mutation screening strategy was established for simi lar to 80% of the genomic coding sequence (exons 2, 11, 13-16, 20, 24) . This strategy was applied to screen genomic DNA from 50 familial bre ast and/or ovarian cancer patients who had previously been examined fo r BRCA1 mutations by SSCP. A total of 14 carriers of 12 distinct disea se-associated mutations and 7 carriers of 6 distinct rate substitution s leading to amino acid substitutions were identified. The SSCP failed to detect 40% of the different deletions/insertions (4/10) and 75% (6 /8) of the different base substitutions leading to terminating codons or rare amino acid changes, SSCP did, however, identify one rare base substitution that could not be detected in the CDGE screening, To eval uate the CDGE mutation screening strategy further, 25 unrelated patien ts from Norwegian breast and/or ovarian cancer families were examined for BRCA1 mutations using a combined genomic DNA/cDNA approach coverin g the entire coding sequence of the gene, A total of six mutation carr iers were detected, all of whom had cases of ovarian cancer in their f amilies. Three patients from independent families carried an 1135insA mutation in exon 11, two others had a Gly484ter and an 1675delA mutati on, respectively, and the sixth carried a splice mutation (5194-2 a--> c) causing deletion of exon 18. CDGE may become an efficient tool in d iagnostic and population based screening for BRCA1 mutations. (C) 1998 Wiley-Liss, Inc.