Patent ID: 9506115
Filing Date: 2016-11-29
CPC Classification: C12Q

Claim Text:
1. A method of detecting the absence of a wild type allele of a locus in the JAK2 gene having at least first mutant and wild type alleles, the method comprising: a) providing a reaction mix comprising b) maintaining the reaction mix at a temperature between the probe:mutant allele Tm and the probe:wild type allele Tm, such that the probe preferentially hybridises to the wild type allele; c) carrying out a thermal cycling amplification on the reaction mix, the amplification including a melt phase, an annealing phase, and an extension phase, in which the temperatures of the extension and annealing phases are between the probe:mutant allele Tm and the probe:wild type allele Tm, such that the probe is hybridised to the wild type allele during these phases; to thereby amplify the mutant allele; and d) detecting hybridisation of the probe to the sample at a temperature at or below the probe:mutant allele Tm; detecting hybridisation of the probe to the sample at a higher temperature at or below the probe:wild type allele Tm; and comparing the two; to thereby detect the absence of the wild type allele, wherein if there is hybridisation of the probe to the sample at a temperature at or below the probe:mutant allele Tm and higher than the probe:wild-type allele Tm but not at a temperature higher than the probe:mutant allele Tm, then the wild type allele is absent; and wherein one or both of the primers has a binding site such that the probe competes with the primer for binding, to thereby prevent binding of the primer and hence strand extension.