Evaluation of a novel reverse-hybridization StripAssay for typing DNA variants useful in diagnosis of adult-type hypolactasia

BackgroundAdult-type hypolactasia is a genetically determined inability to digest lactose after weaning. Two single-nucleotide polymorphisms (C-13910T, G-22018A) located upstream of the lactase gene (LCT) within the gene MCM6 are associated with the lactase persistence/non-persistence trait in patients of European descent. Therefore, the genotyping of these SNPs has been established as a diagnostic tool for adult-type hypolactasia. We have recently shown that several novel allelic variants located in close proximity to the C-13910T SNP interfere with the diagnostic accuracy of real-time PCR-based genotyping methods.MethodsWe describe here the validation of a comprehensive reverse-hybridization teststrip-based assay for the detection of common and novel LCT SNPs (C-13907G, C-13910T, T-13913C, G-13914A, T-13915G, and G-22018A). This assay is based on multiplex DNA amplification and ready-to-use membrane teststrips containing variant-specific oligonucleotide probes immobilized as an array of parallel lines.ResultsWe evaluated the novel reverse-hybridization StripAssay on 125 DNA samples in comparison to LightCycler analysis and sequencing. The outcome of StripAssay genotyping was found to be completely concordant with that obtained by sequencing.ConclusionsThe StripAssay represents an accurate and robust screening tool to identify multiple LCT/MCM6 variants in a rapid manner. It overcomes diagnostic pitfalls that were reported and allows the simultaneous genotyping of closely spaced LCT variant sites in a single-step diagnostic approach.