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Nonequilibrium Hybridization Enables Discrimination of a Point Mutation within 5–40 °C
- Stancescu, Maria, Fedotova, Tatiana
A., Hooyberghs, Jef, Balaeff, Alexander, Kolpashchikov, Dmitry M.
- Journal of the American Chemical Society 2016 v.138 no.41 pp. 13465-13468
- DNA, RNA, ambient temperature, biotechnology, hybridization, medicine, point mutation, single nucleotide polymorphism
- Detection of point mutations and single nucleotide polymorphisms in DNA and RNA has a growing importance in biology, biotechnology, and medicine. For the application at hand, hybridization assays are often used. Traditionally, they differentiate point mutations only at elevated temperatures (>40 °C) and in narrow intervals (ΔT = 1–10 °C). The current study demonstrates that a specially designed multistranded DNA probe can differentiate point mutations in the range of 5–40 °C. This unprecedentedly broad ambient-temperature range is enabled by a controlled combination of (i) nonequilibrium hybridization conditions and (ii) a mismatch-induced increase of equilibration time in respect to that of a fully matched complex, which we dub “kinetic inversion”.