Patent ID: 9605302
Date: 2017-03-28
CPC Classifications: C12Q

Claim:
1. A method of detecting at least one target nucleotide sequence, the method comprising: amplifying the at least one target nucleotide sequence with one or more primers comprising a 5′ anchor region, a 3′ extension region, and a bubble region therebetween, wherein the bubble region comprises a nuclease recognition sequence for a restriction enzyme and a cleavage site for said restriction enzyme; treating the amplified nucleotide sequence with said restriction enzyme so as to remove the 5′ anchor region and part of the bubble region from the amplified nucleotide sequence, thereby generating a probe, wherein said probe comprises the 3′ extension region and a remaining part of the bubble region; contacting the at least one target nucleotide sequence with the probe under hybridizing conditions; and detecting a signal related to the presence, level, and/or sequence variation of the target nucleotide sequence; wherein the 5′ anchor region is about 15 to 30 bp, is configured to hybridize to a nucleotide sequence outside of the target nucleotide sequence, and is about 70 to 100% complementary to said nucleotide sequence outside of the target nucleotide sequence; the 3′ extension region is about 5 to 30 bp and about 80 to 100% complementary to a portion of the target nucleotide sequence; and the bubble region is about 50% or less complementary to the target nucleotide sequence.