title: A reusable quartz crystal microbalance biosensor for highly specific detection of single-base DNA mutation.
abstract: A reusable quartz crystal microbalance (QCM) biosensor based on the toehold-mediated strand displacement reaction (SDR) is first developed for highly specific detection of single-base DNA mutation. N⁵, N¹⁰-methylenetetrahydrofolate reductase (MTHFR) gene C677T mutation is chosen as a model to investigate the performance of the constructed biosensor. The capture DNA immobilized on the gold electrode surface of QCM hybridizes with the conjugate of biotinylated reporter DNA and streptavidin (abbreviated to reporter probe) to form the recognition layer. A toehold domain in the reporter DNA is specifically designed to bind the complementary mutant DNA and trigger the SDR, eventually resulting in the release of the complex of reporter probe and mutant DNA from the gold surface, and then the recognition layer is mildly regenerated by subsequently injecting the reporter probe again. The fabricated biosensor shows good repeatability for the mutant DNA, as more than 87% signal intensity of the sensor remained after six repetitive measurements. A linear range of 1-75 nM is achieved with a detection limit of 0.8 nM at room temperature. This biosensor also affords remarkable specificity to the mutant DNA against its wild-type DNA with a discrimination factor of 33. The potential of applying this biosensor in screening real samples is confirmed by spiked HeLa cells lysate with a recovery of 108%. The proposed sensing strategy provides a simple and universal solution for repetitive and highly specific detection of single-base DNA mutation.
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.
authors.forename: Dingzhong
authors.initials: D
authors.lastname: Wang
authors.forename: Gengjia
authors.initials: G
authors.lastname: Chen
authors.forename: Huaming
authors.initials: H
authors.lastname: Wang
authors.forename: Wei
authors.initials: W
authors.lastname: Tang
authors.forename: Wei
authors.initials: W
authors.lastname: Pan
authors.forename: Na
authors.initials: N
authors.lastname: Li
authors.forename: Feng
authors.initials: F
authors.lastname: Liu
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D004247:DNA / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D017354:Point Mutation; D058525:Quartz Crystal Microbalance Techniques; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D004247:DNA