Single Nucleotide Polymorphism Genotyping in Single-Molecule Electronic Circuits

Abstract

Establishing low-cost, high-throughput, simple, and accurate single nucleotide polymorphism (SNP) genotyping techniques is beneficial for understanding the intrinsic relationship between individual genetic variations and their biological functions on a genomic scale. Here, a straightforward and reliable single-molecule approach is demonstrated for precise SNP authentication by directly measuring the fluctuations in electrical signals in an electronic circuit, which is fabricated from a high-gain field-effect silicon nanowire decorated with a single hairpin DNA, in the presence of different target DNAs. By simply comparing the proportion difference of a probe-target duplex structure throughout the process, this study implements allele-specific and accurate SNP detection. These results are supported by the statistical analyses of different dynamic parameters such as the mean lifetime and the unwinding probability of the duplex conformation. In comparison with conventional polymerase chain reaction and optical methods, this convenient and label-free method is complementary to existing optical methods and also shows several advantages, such as simple operation and no requirement for fluorescent labeling, thus promising a futuristic route toward the next-generation genotyping technique.

Keywords: label‐free detection; silicon nanowires; single nucleotide polymorphism; single‐molecule devices.

Figure 1

a) Schematic diagram of single‐molecule biosensors, where the interaction between the hairpin probe and the target DNA represents their hybridization process, not implying the conformation of the target binding to the hairpin loop. b) Schematic demonstration of three‐phase transitions during hairpin DNA hybridization with the complementary target. c,d) 5 s interval source–drain current fluctuations ΔI _D(t) of a representative single hairpin DNA‐decorated SiNW biosensor measured in a pure PBS solution and a PBS solution containing 1 × 10⁻⁶ m complementary target (WT‐C DNA) at T = 45 °C, respectively. Insets show representative data over a short time interval. The right panels are the corresponding histograms of current values, revealing c) two and d) three Gaussian peaks in conductance.

Figure 2

Hybridization for allele detection. Source–drain current fluctuations ΔI _D(t) of the representative single hairpin DNA‐decorated SiNW biosensor in the presence of 1 × 10⁻⁶ m mismatched target DNA solution at T = 45 °C (a: MT‐A; b: MT‐G; c: MT‐T). The inserts in left panels are the amplified data over a short time interval. The right panels are the corresponding histograms of the conductance, revealing three Gaussian peaks. The inserts in right panels are the enlarged peaks for the low state.

Figure 3

Distributions of the duration for the three conductance states: a–d) high state, b–e) intermediate state, and c–f) low state, extracted from the hybridization data of a hairpin DNA with WT‐C (above) and MT‐T (bottom), respectively. For each state, two distributions are shown to distinguish the direction of the conformational changes.

Figure 4

Comparisons of hybridization dynamics. a) Distributions of the mean lifetimes of each state. b) Distributions of the transition probability among the three states. c) Percentages of three conductance states of hairpin‐DNA hybridization in the presence of WT‐C, MT‐A, MT‐G, and MT‐T. Error bars were calculated from at least five groups of 5 s interval fluctuating data sets for a well‐matched target and 60 s interval for single‐base mismatched targets.