Patent ID: 9404149
Filing Date: 2016-08-02
CPC Classification: C12Q

Claim Text:
1. A set of primers for detecting, by loop mediated isothermal amplification, the presence of a point mutation in a target nucleic acid molecule in a background of wild type nucleic acid molecules, the set of primers comprising: i. a first outer primer F3 and a second outer primer B3; ii. a first inner primer FIP and a second inner primer BIP, wherein FIP comprises a 3′ nucleic acid sequence F2 and a 5′ nucleic acid sequence F1c and BIP comprises a 3′ nucleic acid sequence B2 and a 5′ nucleic acid sequence B1c, wherein F1c is able to recognize and hybridize to a region of the target nucleic acid molecule designated as F1 and wherein B1c is able to recognize and hybridize to a region of the target nucleic acid molecule designated as B1, and wherein F1 and B1 are different regions located on opposite strands of the target nucleic acid molecule, wherein F2 is complementary to a region of the target nucleic acid molecule designated as F2c and B2 is complementary to a region of the target nucleic acid molecule designated as B2c, wherein F2c and B2c are non-overlapping regions located on opposite strands of the target nucleic acid molecule, wherein either B2c is downstream of the point mutation or F2c is upstream of the point mutation, and wherein if B2c is downstream of the point mutation, then said point mutation is located in the F2c sequence or between the F2c sequence and the F1c sequence, or if F2c is upstream of the point mutation, then said point mutation is located in the B2c sequence or between the B2c sequence and the B1c sequence; iii. a stem-loop mutant extensible primer comprising: a central loop sequence complementary to a region of a target nucleic acid molecule comprising the point mutation, and a 5′ end sequence and a 3′ end sequence which are complementary to each other such as to form a stem upon intramolecular hybridization, the hybridization affinity of the central loop sequence to the region of the target nucleic acid molecule comprising the point mutation being higher than the intramolecular hybridization affinity of the 5′ sequence to the 3′ sequence, and a non-extensible moiety capable of selectively hybridizing to the wild type nucleic acid molecules and of blocking amplification thereof.