Method of detecting a polymorphism at a polymorphism site

Abstract

The present invention provides a method for detecting a mutation capable of detecting a mutation with high sensitivity and high reliability in one reaction system. Using primers (Xmt) and (Xwt), a target nucleic acid sequence whose objective base to be detected is a mutant-type is amplified with amplification efficiency higher than a target nucleic acid sequence whose objective base to be detected is a normal-type. The (Xmt) is a primer that is complementary to a region including a mutant-type base in the template nucleic acid and has a base complementary to a mutant-type base at a 3' region, and the (Xwt) is a primer that is complementary to a region including a normal-type base in the template nucleic acid and has a base complementary to a normal-type base at a 3′ region. It is preferable that amplification efficiency by the (Xmt) with reference to a mutant-type template nucleic acid is higher than that by the (Xwt) with reference to a normal-type template nucleic acid. Then, a signal value that shows a molten state of a hybridization product between the thus obtained amplification product and the probe is measured, and the presence or absence of the mutation of the objective base site is determined from a change in the signal value accompanying a change in the temperature.