Description and validation of high-throughput simultaneous genotyping and mutation scanning by high-resolution melting curve analysis

Tú Nguyen-Dumont, Florence Le Calvez-Kelm, Nathalie Forey, Sandrine McKay-Chopin, Sonia Garritano, Lydie Gioia-Patricola, Deepika De Silva, Ron Weigel, Suleeporn Sangrajrang, Fabienne Lesueur, Sean V. Tavtigian

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35 Citations (Scopus)


Mutation scanning using high-resolution melting curve analysis (HR-melt) is an effective and sensitive method to detect sequence variations. However, the presence of a common SNP within a mutation scanning amplicon may considerably complicate the interpretation of results and increase the number of samples flagged for sequencing by interfering with the clustering of samples according to melting profiles. A protocol describing simultaneous high-resolution gene scanning and genotyping has been reported. Here, we show that it can improve the sensitivity and the efficiency of large-scale case-control mutation screening. Two exons of ATM, both containing an SNP interfering with standard mutation scanning, were selected for screening of 1,356 subjects from an international breast cancer genetics study. Asymmetric PCR was performed in the presence of an SNP-specific unlabeled probe. Stratification of the samples according to their probe-target melting was aided by customized HR-melt software. This approach improved identification of rare known and unknown variants, while dramatically reducing the sequencing effort. It even allowed genotyping of tandem SNPs using a single probe. Hence, HR-melt is a rapid, efficient, and cost-effective tool that can be used for high-throughput mutation screening for research, as well as for molecular diagnostic and clinical purposes.

Original languageEnglish
Pages (from-to)884-890
Number of pages7
JournalHuman Mutation
Issue number6
Publication statusPublished - Jun 2009
Externally publishedYes


  • ATM
  • Genotyping
  • High throughput mutation scanning
  • High-resolution melting curve analysis
  • HR-melt

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