title: Advantage of high-resolution melting curve analysis over conformation-sensitive gel electrophoresis for mutational screening of BRCA1 and BRCA2 genes.
abstract: BACKGROUND
  Mutation screening of BRCA1 and BRCA2 (BRCAs) genes is a time-consuming and costly procedure that demands faster and cheaper alternative methods for routine diagnostics. The present study is aimed at comparing the results obtained with screening mutations methods, conformation sensitive gel electrophoresis (CSGE) and high-resolution melting analysis (HRMA), for BRCAs attending to their specificity, sensitivity, reliability and cost-efficiency.
  
  
  METHODS
  We included 52 DNA samples of index patients from high-risk families. The mutational screening was performed by CSGE according to the Ganguly (1993) method and HRMA according to a modified De Leeneer (2008) method. The assays were performed in 384 well plates in the LightCycler 480 (Roche). All PCR products showing altered patterns were confirmed by sequencing.
  
  
  RESULTS
  The results obtained with the mutational study of BRCAs genes showed that HRMA exhibited higher sensitivity than CSGE as it was able to detect a wide mutational spectra of genetic variants in a larger number of samples. Aditionally, the combination of HRMA with hybiridization probes in a second step of the assay allows the specific confirmation of mutations. Furthermore, HRMA use less time, allowing the reduction of analysis time.
  
  
  CONCLUSIONS
  HRMA offers clear advantages over CSGE for the mutation screening of BRCAs genes as it has greater sensitivity and higher efficiency and it is less time-consuming.
authors.affiliation: Laboratory of Molecular Biology, Clinical Analysis Service, Hospital Universitario La Fe, Valencia, Spain.
authors.forename: Inmaculada
authors.initials: I
authors.lastname: de Juan Jiménez
authors.forename: Eva Esteban
authors.initials: EE
authors.lastname: Cardeñosa
authors.forename: Sarai Palanca
authors.initials: SP
authors.lastname: Suela
authors.forename: Eva Barragán
authors.initials: EB
authors.lastname: González
authors.forename: Dolores Salas
authors.initials: DS
authors.lastname: Trejo
authors.forename: Oscar Fuster
authors.initials: OF
authors.lastname: Lluch
authors.forename: Pascual Bolufer
authors.initials: PB
authors.lastname: Gilabert
mesh_terms: D001483:Base Sequence; D015342:DNA Probes; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D019398:Genes, BRCA1; D024522:Genes, BRCA2; D005820:Genetic Testing; D006801:Humans; D009154:Mutation; D016133:Polymerase Chain Reaction
chemical_list: D015342:DNA Probes