A novel SYBR-based duplex qPCR for the detection of gene dosage: detection of an APC large deletion in a familial adenomatous polyposis patient with an unusual phenotype

Author(s)

Torrezan, Giovana Tardin ^[1] ; Carneiro da Silva, Felipe Cavalcanti ^[1] ; Victorino Krepischi, Ana Cristina ^{[2, 1]} ; Monteiro dos Santos, Erika Maria ^[1] ; Rossi, Benedito Mauro ^[3] ; Carraro, Dirce Maria ^{[2, 1]}

Affiliation

^[1] AC Camargo Hosp, CIPE, Int Ctr Res & Training, BR-01508010 Sao Paulo - Brazil

^[2] Natl Inst Sci & Technol Oncogen INCITO, Sao Paulo - Brazil

^[3] Barretos Canc Hosp, Pio Fdn 12, Barretos, SP - Brazil

• Document type: Journal article
• Source: BMC MEDICAL GENETICS; v. 13, JUL 16 2012.
• Web of Science Citations: 11

Abstract

Background: Familial adenomatous polyposis (FAP) is a hereditary colorectal cancer syndrome caused by a loss of function of the APC gene. Large deletions in APC are a common cause of FAP; despite the existence of a variety of gene dosage detection methodologies, most are labor intensive and time and resource consuming. Methods: We describe a new duplex qPCR method for gene dosage analysis based on the coamplification of a target and a reference gene in a SYBR Green reaction, followed by a comparison of the ratio between the target and the reference peaks of the melting curve for the test (patient) and control samples. The reliability of the described duplex qPCR was validated for several genes (APC, HPRT1, ATM, PTEN and BRCA1). Results: Using this novel gene dosage method, we have identified an APC gene deletion in a FAP patient undergoing genetic testing. Comparative genomic hybridization based on microarrays (aCGH) was used to confirm and map the extent of the deletion, revealing a 5.2 MB rearrangement (5q21.3-q22.3) encompassing the entire APC and 19 additional genes. Conclusion: The novel assay accurately detected losses and gains of one copy of the target sequences, representing a reliable and flexible alternative to other gene dosage techniques. In addition, we described a FAP patient harboring a gross deletion at 5q21.3-q22.3 with an unusual phenotype of the absence of mental impairment and dysmorphic features. (AU)

• FAPESP's process: 08/57887-9 - National Institute of Oncogenomics
• Grantee: Luiz Paulo Kowalski
• Support Opportunities: Research Projects - Thematic Grants