Genetic diseases. Development of a rapid protocol for mutation detection by microarray analysis

Several genetic diseases are characterized by genes with one or few very frequent mutations. Here we report the development of a new protocol based on arrayed primer extension (APEX) technique for the detection of very common alleles. In this new protocol, named arrayed amplicon primer extension (AAPEX), for the first time instead of linking amino-modified oligonucleotides on the glass surface, we have attached on the same surface PCR products corresponding to different patients and carriers. These products have been obtained using 1 amino-modified oligo out of the 2 PCR primers. This protocol has been developed for the detection of 1 of the 2 most common alleles (H63D) of HFE gene, which is mutated in hereditary hemochromatosis, one of the most common recessive diseases in Caucasians. A series of experiments performed with this protocol allowed us to demonstrate that it can be easily applied to the simultaneous screening of several individuals. Our results clearly demonstrate the feasibility, reliability and specificity of this protocol for a mutational screening of a large series of individuals.