Pitfalls in genetic testing: a case of a SNP in primer-annealing region leading to allele dropout in BRCA1

Autor(es)

Silva, Felipe Carneiro ; Torrezan, Giovana Tardin ; Brianese, Rafael Canfield ; Stabellini, Raquel ; Carraro, Dirce Maria

• Tipo de documento: Artigo Científico
• Fonte: MOLECULAR GENETICS & GENOMIC MEDICINE; v. 5, n. 4, p. 443-447, JUL 2017.
• Citações Web of Science: 4

Resumo

Background Hereditary breast and ovarian cancer is characterized by mutations in BRCA1 or BRCA2 genes and PCR-based screening techniques, such as capillary sequencing and next-generation sequencing (NGS), are considered gold standard methods for detection of pathogenic mutations in these genes. Single-nucleotide polymorphisms (SNPs) constitute a vast source of variation in the human genome and represent a risk for misdiagnosis in genetic testing, since the presence of a SNP in primer-annealing sites may cause false negative results due to allele dropout. However, few reports are available and the frequency of this phenomenon in diagnostic assays remains unknown. Methods and Results In this article, we investigated the causes of a false negative capillary sequencing result in BRCA1 involving a mother-daughter dyad. Using several molecular strategies, including different DNA polymerases, primer redesign, allele-specific PCR and NGS, we established that the initial misdiagnosis was caused by a SNP located in the primer-annealing region, leading to allele dropout of the mutated allele. Conclusion Assuming that this problem can also occur in any PCR-based method that are widely used in diagnostic settings, the clinical report presented here draws attention for one of the limitations of genetic testing in general, for which medical and laboratory communities need to be aware. (AU)

• Processo FAPESP: 13/23277-8 - Aspectos moleculares envolvidos no risco, desenvolvimento e progressão do carcinoma ductal de mama: busca de novos genes de susceptibilidade e investigação da progressão do carcinoma in situ e do papel da mutação em BRCA1 no tumor triplo negativo
• Beneficiário: Dirce Maria Carraro
• Modalidade de apoio: Auxílio à Pesquisa - Temático