Construction Of DNA Machines And Discrimination Of Single-Base Changes On Chip Surface

Genetic variations with single-base changes, such as insertions, deletions, and single-base mismatch, are closely associated with physical and genetic diseases among humans. The genotyping of genetic variations in human DNA is relevant to early diagnosis of diseases, investigation of human response to drugs, and prevention and treatment of specific human disease, Therefore, a simple, accurate, parallel, and time-saving method must be developed, to detect nucleic acid sequences that have a single-base resolution. We presented a new single-base change detection method based on enhanced toehold exchange on chip surface. Our experimental results proved and demonstrated the capacity of the toehold exchange probe to differentiate DNA chains with single-base variations. The toehold exchange reaction proceeded more efficiently on chip surface (86.1%) than in solution (24.9%). We examined the method on single-base mutations, insertions, and deletions at different positions along a DNA chain. With this method, we clearly discriminated DNA strands with single-base changes at arbitrary positions with nucleic acid concentrations ranging from0.8nM to1μM. Thus, the detection limit is0.8nM. Dual-polarization interferometry (DPI) was used to monitor the entire process, and the efficiency was calculated on the basis of the mass changes on the surface. The method is simple, versatile, and capable of detecting single-base changes in both the toehold and the double-stranded DNA regions. A simple DNA microarray method was also developed to detect single-base changes.Meanwhile, DNA has been demonstrated to be an extremely versatile building material for the assembly of novel nanostructures and nanodevices with unique functions. In recent years, DNA-based nanomachines have been shown to be capable of producing reversible, well-defined nanometer-scale motions. We have implemented DNA machine on a new type of microfluidic DNA microarray chip, which combined the advantages of microfluidics and DNA microarrays. Basing on the two-step toehold exchange reaction, we realized fluorescent signal amplification to succeed in building signal amplifying DNA sensors. We also constructed "AND" logic gates on the surface of microfluidic DNA microarray chips.At last, microfluidics is a novel technology that deals with the precise control and manipulation of small-volume fluids in channels that are less than a millimeter. We conducted experiments on uniquely designed microfluidic chips that generate droplets through a microfluidic flow-focusing approach. The fluid flow in the microfluidic channel produced a shear flow field at low Reynolds numbers. The droplets in the microfluidic system exhibited special droplet pattern formations similar to periodic crystal-like lattices because of the competition between shear forces and surface tension. Considering that geometric effects and relative fluid are significant in pattern formation, we studied two types of emulsification by adjusting the flow rate ratio Qi/Qo of water (droplet phase) and oil (continuous phase) and by changing the outlet channel widths. We observed droplet patterns that range from monolayer dispersion to multilayer arrangement and monolayer squeezing arrangement. Our results provide a new detailed strategy to control drop size and drop pattern formation in emulsions in a microfluidic system. Our strategy may serve as a theoretical guidance to synthesize dispersed or polydispersed colloidal particles.