title: Determination of mutated genes in the presence of wild-type DNA by using molecular beacons as probe.
abstract: Low-abundance mutations in the presence of wild-type DNA can be determined using molecular beacon (MB) as probe. A MB is generally used as DNA probe because it can distinguish single-base mismatched target DNA from fully matched target DNA. However, the probe can not determine low-abundance mutations in the presence of wild-type DNA. In this study, this limitation is addressed by enhancing the stability of unpaired base-containing dsDNA with a hydrogen-bonding ligand, which was added after hybridization of the MB to the target DNA. The ligand formed hydrogen bonds with unpaired bases and stabilized the unpaired base-containing dsDNA if target DNA is mutated one. As a result, more MBs were opened by the mutant genes in the presence of the ligand and a further increase in the fluorescence intensity was obtained. By contrast, fluorescence intensity did not change if target DNA is wild-type one. Consequent increase in the fluorescence intensity of the MB was regarded as a signal derived from mutant genes. The proposed method was applied in synthetic template systems to determine point mutation in DNA obtained from PCR analysis. The method also allows rapid and simple discrimination of a signal if it is originated in the presence of mutant gene or alternatively by a lower concentration of wild gene.
authors.affiliation: Key Laboratory of Analytical Chemistry for Life Science of Shaanxi Province, School of Chemistry and Chemical Engineering, Shaanxi Normal University, Xi'an 710062, China; School of Chemistry and Chemical Engineer, Luoyang Normal University, Luoyang 471022, China.
authors.forename: Yonghua
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Key Laboratory of Analytical Chemistry for Life Science of Shaanxi Province, School of Chemistry and Chemical Engineering, Shaanxi Normal University, Xi'an 710062, China.
authors.forename: Junjie
authors.initials: J
authors.lastname: Ai
authors.affiliation: Key Laboratory of Analytical Chemistry for Life Science of Shaanxi Province, School of Chemistry and Chemical Engineering, Shaanxi Normal University, Xi'an 710062, China.
authors.forename: Qiaorong
authors.initials: Q
authors.lastname: Gu
authors.affiliation: Key Laboratory of Analytical Chemistry for Life Science of Shaanxi Province, School of Chemistry and Chemical Engineering, Shaanxi Normal University, Xi'an 710062, China. Electronic address: gaoqiang@snnu.edu.cn.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Gao
authors.affiliation: Key Laboratory of Analytical Chemistry for Life Science of Shaanxi Province, School of Chemistry and Chemical Engineering, Shaanxi Normal University, Xi'an 710062, China.
authors.forename: Honglan
authors.initials: H
authors.lastname: Qi
authors.affiliation: Key Laboratory of Analytical Chemistry for Life Science of Shaanxi Province, School of Chemistry and Chemical Engineering, Shaanxi Normal University, Xi'an 710062, China.
authors.forename: Chengxiao
authors.initials: C
authors.lastname: Zhang
mesh_terms: D002942:Circular Dichroism; D004247:DNA / Q000235:genetics*; D005796:Genes; D015335:Molecular Probes / Q000737:chemistry*; D009154:Mutation / Q000235:genetics*; D009691:Nucleic Acid Denaturation; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D013050:Spectrometry, Fluorescence; D013696:Temperature
chemical_list: D015335:Molecular Probes; D004247:DNA
keywords: DNA; Determinate; Hydrogen bonding; Low-abundance mutation; Molecular beacon