| 主权项 |
1. A method of detecting a target nucleic acid in a sample, the method comprising
(a) incubating in a reaction mixture:
i) a sample comprising a target nucleic acid; andii) a pair of proximity probes comprising a first and second probe, where:a first probe comprises a target binding (TB) segment that hybridizes to a first target (T) segment of the target nucleic acid, and an interacting (I) segment at the 3′ end of the probe, wherein the I segment is complementary to an I segment at the 3′ end of the second probe; anda second probe comprises a TB segment that hybridizes to a second, non-overlapping T segment of the target nucleic acid that is in close proximity to the first T segment, and an I segment at the 3′ end, wherein the 3′ sequence is complementary to the I segment of the first probe, wherein the reaction mixture is incubated under conditions in which the TB segment of the first probe hybridizes to the first T segment of the target nucleic acid and the TB segment of the second probe hybridizes to the second T segment of the target nucleic acid, thereby allowing the I segment of the first probe to hybridize to the I segment of the second probe to form a duplex comprising the I segments of the first and second probe; (b) adding a DNA polymerase and maintaining the reaction mixture under conditions in which the first and/or second probe is extended to obtain a first extended product; (c) amplifying the extended product, or a subregion thereof in an amplification reaction mixture comprising a pair of amplification primers that amplify the first extended product, or subregion thereof; (d) detecting the amplicon obtained in (c). |
