Recognition of single nucleotide polymorphisms using scanning potential hairpin denaturation.
Conventional single nucleotide polymorphism (SNP) assays, which based their detection on the stringency or temperature of the washing buffers, have encountered difficulties to distinguish a single base pair mismatch from a perfect match. In this study, scanning potential hairpin denaturation (SPHD) has been developed to detect SNP in a sensitive and reliable manner. Combined with hairpin oligonucleotide probes, scanning surface electric potential was used to induce a dissociation of double-stranded DNA around a unique "melting potential" (Vm), and it generated a high-contrast SNP recognition signal. A 21 base pair p53 gene segment was used to test this novel method. A single nucleotide mismatch to the hairpin probes caused an average of 400-800 mV difference in melting potential against the perfect match, while the error of this assay was lower than 20 mV. Experiments demonstrated that the hairpin stem was critical to the method. The concept of scanning potential hairpin denaturation could also be used extensively in different areas of nucleotide hybridization based assays.
Wei
Fang
F
State Key Laboratory of Molecular Dynamic and Stable Structures, and Department of Chemical Biology, College of Chemistry, Peking University, Beijing 100871, China.
Chen
Chunlai
C
Zhai
Lin
L
Zhang
Ning
N
Zhao
Xin Sheng
XS
D004247:DNA / Q000737:chemistry* / Q000235:genetics*; D015342:DNA Probes; D004563:Electrochemistry; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization; D020641:Polymorphism, Single Nucleotide; D013499:Surface Properties; D013816:Thermodynamics
D015342:DNA Probes; D004247:DNA