Genotyping of frequent BRCA1/2 SNPs with unlabeled probes: a supplement to HRMCA mutation scanning, allowing the strong reduction of sequencing burden.

ABSTRACT: We previously validated mutation scanning for BRCA1 and 2 using high-resolution melting curve analysis (HRMCA). Due to recurrent single nucleotide polymorphisms (SNPs), a considerable amount of sequencing work remains after HRMCA, as melting curves for SNPs and deleterious mutations may be similar. Here, we present a simple approach for the optimization of SNP genotyping with HRMCA using unlabeled probes. Protocols were optimized for 14 frequent SNPs in BRCA1 and 2. Two probes contained an additional mismatch to detect a rare polymorphism a few nucleotides upstream. PCR was performed in the presence of LCgreenPlus and analyzed on a Lightscanner. Genotyping assays were optimized with five wild-type, heterozygous, and homozygous mutant samples. Sensitivity and specificity of the assays were

SUBMITTER: De Leeneer K 

PROVIDER: S-EPMC2729838 | biostudies-literature | 2009 Sep

REPOSITORIES: biostudies-literature