Abstract: The inventions deals with determination of the human hemochromatosis gene (HFE) mutation (C282Y of HFE protein), responsible for the disease of hereditary hemochromatosis. The invention is important in diagnosis and risk assessment for this disease. The method consists of a single-tube high-throughput PCR assay for the detection of C282Y. We invented that it is advantageously possible to combine three concepts each known separately in prior art from different sources: allele specific PCR, mutagenically separated PCR, and amplicon identification by specific dissociation curves. Analysis can be performed in either a conventional or fluorescence-detecting thermocycler using the same primers, reactant constituents and cycling protocol. PCR products are identified either by their length or melting temperature (Tm). Primer cross reactions are prevented by deliberate primer: primer and primer: template mismatches.