Design And Fabrication Of Solid-state Nanopore Sensors For Single Molecules Detection

The research field of nanopore based sensors has developed rapidly within the past two decades, and has become widely accepted. One of the reasons for the popularity is the simplicity of the approach. To data, nanopore based sensing is the most promising technology for single molecules analytics. The range of analytes that can be detected with nanopores currently includes small molecules, organic polymers, peptides, proteins, enzymes, and biomolecular complexes. More importantly, it has the potential to sequence DNA at high speed and low cost. In this dissertation,we developed a controllable solid-state nanopore fabrication method; investigated the effects of electrolyte temperature on DNA molecule translocation; and studied the ion transport through nanopores under the condition of high ionic strength and an electrolyte concentration gradient across the nanopore. The key results are summarized as below:(1) Si3N4 and Al2O3 nanopore sensors can be fabricated with desirable diameter and length. The nanopore fabrication process includes two steps. Firstly, we designed a micro-electro-mechanical systems (MEMS) process to fabricate the silicon chips with free-standing Si3N4 membrane. The second step is to drill a nanopore on the Si3N4 membrane by the means of focus ion beam (FIB) or transmission electron microscopy (TEM). We investigated the relation between fabrication parameters and pore dimensions. The Si3N4 nanopores with diameter less than 2 nm and length less than 20 nm can be fabricated by a strategy, which is milling the Si3N4 membrane to reduce its thickness firstly and then drilling a pore in the milled region. We also proposed a novel Al2O3 nanopore manufacture process with' controllable size, based on atomic layer deposition (ALD), and can be expanded to almost all the materials deposited by ALD. The Si3N4 and Al2O3 nanopore sensors were used to detect DNA molecules. Results show that blockade current and translocation time are similar for both Al2O3 and Si3N4 nanopore with similar size. But the Al2O3 nanopore sensors demonstrate lower 1/f noise than Si3N4 nanopore sensors, thus with better detection properties.(2) In order to slow down the translocating molecules, we systemically studied the effects of electrolyte temperature on DNA molecule translocation. Lower temperature leads to the slowed DNA translocation speed, which stems from the increased electrolyte viscosity.Furthermore, the DNA translocation speed can be slowed down as long as one side temperature is lowered, regardless of the temperature gradient direction. This indicates that the thermophoretic driving force generated by a temperature gradient has no obvious effect on the threading speed of DNA molecules. Interestingly, the capture rate of DNA molecules is increased when the temperature gradient direction is same as that of DNA molecules motion. The translocation time and capture rate during DNA translocation from hot side at 21? to cold one at 2? are 2.26 ms and 1.23 s-1 respectively, which are 1.5 times and 1.7 times larger than that under the condition of both chambers at 20?. Therefore, an optimized nanopore detection configuration is proposed to achieve higher capture rates and lower DNA translocation speeds.(3) We seek to explore ionic current modulation when DNA molecules translocate through different size nanopores in high concentration electrolytes and with a concentration gradient across the nanopore. Contrary to our expectation, we observed the ionic current enhancement from DNA translocation even in a rather high concentration range. We further conducted numerical modeling to analyze the underlying mechanisms for this interesting ionic current modulation pattern. Different from the previous understanding that ionic current enhancement is due to the additional ions brought into the nanopore within the electric double layer of the DNA, our analysis indicates that in this case, the enhancement is mainly due to the electroosmotic flow (EOF). With a DNA molecule inside the nanopore, the ions are pumped in from the trans side because of the stronger EOF induced by the surface charge of the DNA. Systematic variation of the pore size and the observed ionic current modulation further confirm our analysis. Smaller diameter nanopores will need larger electrolyte concentration difference to convert the pulse signal from negative to positive for dsDNA translocation events due to EOF rate limitation. This research work complements the theory of ions transport in nanopore.(4) We carried out two series of experiments of DNA molecules detection by Si3N4 nanopore based sensors. Based on the signal analytics and information extraction, we can distinguish different types of DNA molecules, including poly(dT)50, poly(dT)80 and ?-DNA.It is demonstrated that the nanopore based sensors have very high accuracy in single molecule detection. We also identified the configuration of DNA in KCI or Mg2Cl solution by Si3N4 nanopore based sensors. Experimental results indicate that long DNA strands will not attract each other in KCl solution, while the presence of the Mg2+ ions induces the attraction force between the neighboring nucleic acid in long DNA strands. In Mg2C1 solution,long DNA strands will be bridged by Mg2+, resulting in the formation of condensed state.