Patent ID: 9284603
Filing Date: 2016-03-15
CPC Classification: C12Q

Claim Text:
1. A method for detecting a polymorphism in a biological sample as an assay for detecting diseases associated with gene mutations, the method comprising the steps of: amplifying a target sequence in a template nucleic acid in a reaction system containing primers (X1) and (X2), and detecting a polymorphism at a target site in the target sequence with a probe that hybridizes to the target sequence, wherein the target sequence comprises the polymorphic target site, and a base (x) at the target site is either a first base (x1) or a second base (x2), wherein: primer (X1) is a primer comprising a sequence (A1′) and a sequence (E1), the sequence (A1′) being complementary to a partial sequence (A1) in the template nucleic acid, and having, in its 3′ region, a base (x1′) complementary to the first base (x1) at the target site in a 5′ region of the partial sequence (A1), the sequence (E1) being noncomplementary to a partial sequence (B1) adjacent to a 3′ end of the partial sequence (A1) in the template nucleic acid, and being bound to a 5′ end of the sequence (A1′), wherein the number of bases in the sequence (E1) is more than 0 but not more than 5; and primer (X2) is a primer comprising a sequence (A2′) and a sequence (E2), the sequence (A2′) being complementary to a partial sequence (A2) in the template nucleic acid, and having, in its 3′ region, a base (x2′) complementary to the second base (x2) at the target site in a 5′ region of the partial sequence (A2), the sequence (E2) being noncomplementary to a partial sequence (B2) adjacent to a 3′ end of the partial sequence (A2) in the template nucleic acid, and being bound to a 5′ end of the sequence (A2′), wherein the sequence (E2) is different from the sequence (E1) and wherein (E2) has the same base length as (E1), wherein a Tm value of the sequence A2′) of the primer (X2) is higher than a Tm value the sequence (A1′) of the primer (X1).