title: Highly selective detection of single-nucleotide polymorphisms using a quartz crystal microbalance biosensor based on the toehold-mediated strand displacement reaction.
abstract: Toehold-mediated strand displacement reaction (SDR) is first introduced to develop a simple quartz crystal microbalance (QCM) biosensor without an enzyme or label at normal temperature for highly selective and sensitive detection of single-nucleotide polymorphism (SNP) in the p53 tumor suppressor gene. A hairpin capture probe with an external toehold is designed and immobilized on the gold electrode surface of QCM. A successive SDR is initiated by the target sequence hybridization with the toehold domain and ends with the unfolding of the capture probe. Finally, the open-loop capture probe hybridizes with the streptavidin-coupled reporter probe as an efficient mass amplifier to enhance the QCM signal. The proposed biosensor displays remarkable specificity to target the p53 gene fragment against single-base mutant sequences (e.g., the largest discrimination factor is 63 to C-C mismatch) and high sensitivity with the detection limit of 0.3 nM at 20 °C. As the crucial component of the fabricated biosensor for providing the high discrimination capability, the design rationale of the capture probe is further verified by fluorescence sensing and atomic force microscopy imaging. Additionally, a recovery of 84.1% is obtained when detecting the target sequence in spiked HeLa cells lysate, demonstrating the feasibility of employing this biosensor in detecting SNPs in biological samples.
authors: Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.
authors: Dingzhong
authors: D
authors: Wang
authors: Wei
authors: W
authors: Tang
authors: Xiaojie
authors: X
authors: Wu
authors: Xinyi
authors: X
authors: Wang
authors: Gengjia
authors: G
authors: Chen
authors: Qiang
authors: Q
authors: Chen
authors: Na
authors: N
authors: Li
authors: Feng
authors: F
authors: Liu
mesh_terms: D001483:Base Sequence; D015374:Biosensing Techniques / Q000379:methods*; D015342:DNA Probes / Q000737:chemistry* / Q000235:genetics; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000235:genetics; D004566:Electrodes; D006046:Gold / Q000737:chemistry; D006367:HeLa Cells; D006801:Humans; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000235:genetics; D020641:Polymorphism, Single Nucleotide; D058525:Quartz Crystal Microbalance Techniques / Q000379:methods*; D013696:Temperature; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
chemical_list: D015342:DNA Probes; D004277:DNA, Single-Stranded; D009838:Oligodeoxyribonucleotides; D016159:Tumor Suppressor Protein p53; D006046:Gold