High-resolution melting analysis using unlabeled probe and amplicon scanning simultaneously detects several lactase persistence variants.

Lactase persistence and thereby tolerance to lactose is a common trait in people of Northern European descent. It is linked to the LCT −13910C>T variant located in intron 13 of the MCM6 gene 13.9 kb upstream of the lactase ( LCT) gene. In people of African and Middle Eastern descent, lactase persistence can be associated with other variants nearby the −13910C>T variant, limiting the use of the −13910C>T-based SNP analysis, e.g. TaqMan assays for the diagnosis of lactose intolerance. Using high-resolution melting analysis, we identified five samples that were heterozygous for the −13915T>G variant among 78 patients genotyped as −13910C/C by a TaqMan assay. All samples originated from patients of probable Middle Eastern descent. In order to detect the −13910 and −13915 variants simultaneously, we developed a new high-resolution melting (HRM) analysis assay based on unlabeled probe genotyping and simultaneous amplicon scanning analysis. By using this assay we were able to distinguish the −13910 and −13915 genotypes clearly. Furthermore, we identified two rare variants, the −13907C>G and −13913T>C. With this method, based on an inexpensive unlabeled probe, it is possible to simultaneously detect the −13910C>T and −13915T>G variants in addition to rarer variants surrounding the −13910 site. This new method may contribute to improve the diagnostic performance of the genetic analysis for lactose intolerance. [ABSTRACT FROM AUTHOR]