High-resolution melting analysis for genotyping Duffy blood group antigens

Antigens of the Duffy (Fy) blood group are significant in medical transfusions since they may cause serious post-transfusion reactions and hemolytic disease of the fetus and newborn. Results of serotyping performed on donors with reduced or abolished erythrocyte Duffy expression may be misleading, since the Duffy antigen is also present on non-erythroid cells. In such cases only DNA-based genotyping may reveal the actual Duffy antigen status. Here we describe the high-resolution melting (HRM) method for Duffy genotyping, which is a new post-PCR analysis method used for identifying genetic variations in nucleic acid sequences. It is based on the PCR melting curve technique where single nucleotide polymorphism (SNP) in DNA determines a characteristic shape of the melting curve and melting temperature (Tm) of a sample. HRM analysis for FY genotyping can discriminate SNPs in the FY gene through detection of small differences in melting profiles of variants when compared to controls. Recently, we have shown the usefulness of HRM analysis in elucidation of the molecular basis of Duffy-negative phenotype in a Polish family and in large-scale Duffy genotyping.