Theoretical Investigation On Designing And Manufacturing Of A Single-molecule Detection Chip Based On Solid-state Nanopores

The nanopore sequencing technology was started from 1990s,it was observed that the nucleic acid translocate through protein nanopore.With the development of protein nanopore,the DNA sequencing have been realize caused by the enzyme.Nowadays nanopores are widely used such as drug delivery,ionpermeation and biomolecule detection etc.With the development of nanopore,it become high throughput and low cost as a single-molecule sensing device.Caused by the high resolution of nanopore,it is used to detect the properties,activities and interaction of dynamic and static of molecular.The planar liposomes,lipid membranes and polymer membranes are usually used to support the biological pores hold into an electrochemical chamber.With a brief introduction,the most popular three biological pores are discussed below:?-Hemolysin channel,Msp A channel and Phi29 connector channel.Solid-state nanopores considered as a versatile alternative to biological nanopores due to their unique properties including ease of modifications,mechanical robustness,well defined geometries and dimension,and compatibility with various optical and electronic measurement.With the development of materials technology,solid-state nanopores have a wide range of applications in molecular detection,such as fabrication of nanofluidic devices,controlling molecular transport,and monitoring protein interactions etc.A lot of solid-state nanopore fabrication methods have been developed,in which the most popular method is electron beam transmission and focused ion beam sculpting fabrication.Silicon dioxide and silicon nitride membranes have been used several years as substrates caused by their high chemical stability and low mechanical stress.The key result in this paper are summarized as below:First of all,the low noise solid-state nanopore chip was studied by MEMS technology,the RIE etching and FIB etching method were used to reduce the thin chip film.The nanopores were made by TEM and FIB,the pore size and depth can be controlled well.To realize DNA sequencing by the solid-state nanopore,the translocation speed of the DNA in the nanopore should be slowed down to obtain the temporal resolution.In this study,we make a nanopore sensor system integrating an atomic force microscope(AFM)to control the DNA transport.The speed of DNA binding to the probe tip through the nanopore can be controlled by the AFM,the ionic current as well as the force acting on DNA strand while it translocates through a nanopore can simultaneous be measured by the integrated system.The velocity of DNA molecules could be slowed down to 100 nm/s,much less than 1 nt/ms,which has been able to fully meet the requirement for nanopore DNA sequencing.We demonstrate that the current drop and recovery amplitudes step,the force drop and recovery amplitudes step,the capture and release distance when the DNA was capture and release from the nanopore.Further molecular dynamics simulations illustrate the translocation dynamics of DNA when it is pulling through the nanopore.The current blockage and dwell time obtained when the ds DNA translocates through nanopore is accumulated into scatter plots.Ionic current trace recorded at 1000mv as 48kbp ds DNA translocate through 20nm thickness with 35nm alumina nanopore.Here,we apply Schr(?)dinger's first-passage-time distribution formula to study the distribution of DNA translocation time through alumina nanopores.The first-passage-time distribution is solved by production of the Fokker-Plank equation.Two useful parameters are yielded in analyzing the experimental results:the diffusion constant of DNA inside the nanopore and the drift velocity of DNA translocation.By changing the p H value from 5.2 to 10.8 of the electrolyte solution,the drift velocity of DNA translocation and the diffusion constant of DNA inside the nanopore are extremely close almost as 34nm/?s.By changing the p H value of the electrolyte solution,the surface charge density of the wall and the charge of the DNA molecule can be turned,which result in different DNA molecule capture behavior.The capture rate is about 17 s^-1,the DNA molecule translocate through nanopore when the solution p H is 10.8,and 20 s^-1 as the solution p H is 5.2.And a single stranded DNA was detected by a 2nm nanopore,the lasted patch clamp was used to detecting the translocation events.Analysing the effect of the filter frequency and the biased voltage on the nanopore sequencing.Discrimination of single nucleotide by a nanopore remains challenge because of the minor difference among the four type single nucleotides.Here,the blockade currents induced by translocation of single nucleotides through a 1.8 nm diameter silicon nitride nanopore have been measured.It is found that the single nucleotides are driven through the nanopore by an electroosmotic flow instead of electrophoretic force when a bias voltage is applied.The blockade currents for the four types of single nucleotides are unique and differentiable,following the order of the nucleotide volume.Also,the dwell-time for each single nucleotide can last to several hundred microseconds with the advantage of the electroosmotic flow,which is helpful to single nucleotide identification.The dwell-time distributions are found to obey the first-passage time distribution from the 1D Fokker-Planck equation,from which the velocity and diffusion constant of each nucleotide can be deduced.Interestingly,the larger nucleotide is found to translocate faster than the smaller one inside the nanopore because the larger nucleotide has larger surface area that may produce larger drag force induced by the electroosmotic flow,which is validated by molecular dynamics simulations.Nanopore technique allowed us to detect intact viruses and monitor the effect of fs laser on a model virus at the single virus level.Analysis of changes in the ionic current through the nanopore provides information about size and physical properties of viruses before and after laer treatment.