Patent ID: 8460872

Claim:
A method for determining the amount of a minority nucleic acid species in a sample comprising: (a) contacting under amplification conditions a nucleic acid sample comprising a minority species and a majority species, the combination of the minority species and the majority species comprising total nucleic acid in the sample, with: (i) a first set of amplification primers that specifically amplify a first region in sample nucleic acid comprising a feature that (1) is present in the minority nucleic acid species and is not present in the majority nucleic acid species, or (2) is not present in the minority nucleic acid species and is present in the majority nucleic acid species, (ii) a second set of amplification primers that amplify a second region in the sample nucleic acid allowing for a determination of total nucleic acid in the sample, wherein the first region and the second region are different, and (iii) one or more inhibitory oligonucleotides, each comprising a nucleotide sequence complementary to a nucleotide sequence in the second region, wherein the one or more inhibitory oligonucleotides (1) hybridize to the second region but are not extended and (2) reduce the amplification of the second region; thereby generating minority nucleic acid amplification products and total nucleic acid amplification products, wherein the total nucleic acid amplification products are reduced relative to total amplification products that would be generated if no inhibitory oligonucleotide was present, whereby the total nucleic acid amplification products do not dominate the minority nucleic acid amplification products; (b) separating the minority nucleic acid amplification products and total nucleic acid amplification products, thereby generating separated minority nucleic acid amplification products and total nucleic acid amplification products; and (c) determining the fraction of the minority nucleic acid species in the sample based on the amount of each of the separated minority nucleic acid amplification products and total nucleic acid amplification products.