Oligonucleotides for amplification and detection of...

C - Chemistry – Metallurgy – 12 – Q

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C12Q 1/68 (2006.01) C07H 21/00 (2006.01) C12N 15/10 (2006.01) C12P 19/34 (2006.01)

Patent

CA 2305849

The present invention provides oligonucleotides and methods for amplifying, detecting and identifying sequence variations associated with hemochromatosis. It has been found that the reduced efficiency of primer extension by DNA polymerases when the 3' end of a primer does not hybridize perfectly with the target can be adapted for use as a means for distinguishing or identifying the nucleotide in the target which is at the site where the diagnostic mismatch between the detector primer and the target occurs. The efficiency of detector primer extension is detected as an indication of the presence and/or identity of the sequence variation in the target. The inventive methods make use of hemochromatosis gene (HFE)-specific amplification primers and nucleotide mismatches at or near the 3' end of a detector primer to amplify the HFE gene and discriminate between wild-type alleles and single nucleotide polymorphisms which may occur in exon 2 and exon 4 of the (HFE) gene.

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