Experimental And Theoretical Study Of Nanopore Detection Of DNA And Magnetic Beads Binding DNA

Solid state nanopore sequencing is the most promising method to detect DNA sequences at low cost and is the main means and method for the third generation of DNA sequencing.At present,the bottleneck that restricts the sequencing of solid-state nanoscale is that the speed of DNA through the nanopore is too fast under the high voltage electrophoresis drive,which causes the detection technology to be difficult to recognize single base.In view of the above problems,the effects of different buffer concentration and bias voltage on the blocking current and duration of DNA through nanoscale are investigated.The biotin modified DNA are bound to the magnetic beads modified by the modified streptavidin in order to reduce the speed of DNA.The effects of the blocking current and duration of DNA throughing the nanopore are studied at different buffer concentration and bias voltage.The law of DNA and the magnetic bead binding DNA under different concentration and bias voltage is studied using the multi physical field coupling simulation software COMSOL Multiphysics,.The main research methods and results are as follows:1)Using the means of focusing ion beam(FIB),the silicon nitride nanopore that the diameter is of 25 nm which can be used for the detection of DNA,are fabricated by the method of first shearing and then perforation.The theory of DNA and magnetic bead binding DNA through the nanopore and the effect on the pore of DNA are discussed,including the theory of electric double layer,which determines the distribution of ions and the potential of zeta,the theory of electroosmotic flow and and electrophoretic,the theory of dielectric electrophoretic caused by particles,the streptavidinbiotin theorytheory and the patch clamp technique for high precision current detection.2)Explored the experimental methods and steps of the silicon nitride nanopore,explored the effect of different buffer concentration and bias voltage on the DNA through nanopore.In the range of 0.1-1.0mol/L,the blocking current of the DNA increase linearly with the increase of the buffer concentration,while the duration is linearly decreasing and the capture rate decreases gradually.As the bias voltage increases,the duration of the DNA throughing the nanopore decreases,but the decrease is becoming smaller and smaller,(approximate inverse),blocking current increases linearly with the increase of bias voltage,and the acquisition rate also increases linearly.3)The experimental method of magnetic bead binding DNA was investigated.The current characteristic signals of the magnetic bead binding DNA,pure DNA,pure magnetic beads and pure streptavidin and nanoscale were analyzed and different current signals of different substances were obtained.The effects of different buffer concentration and bias voltage on the magnetic bead binding DNA are investigated.It is found that the current blocking signal of DNA bonded by magnetic beads is close to that of pure DNA.However,the duration of the magnetic bead binding DNA increases obviously.With the increase of the buffer concentration,,and the capture rate decreases but the duration increase.With the linear increase of bias voltage,the duration decrease but the reduction increase gradually.This is due to the fact that the distance between streptavidin-biotin linkage is changeless and the acceleration is larger at the beginning stage.The increase of voltage also results in a linear increase of acquisition rate.4)Using the multi physical field coupling simulation software COMSOL Multiphysics to simulate the law of DNA(simplified nanoparticles model)and the magnetic bead binding DNA through the nanopore,the theoretical basis and the specific modeling process are discussed,including the dimensionless substitution,the establishment of a two-dimensional model,the setting of the solution domain and boundary conditions,ODE and boundary variable settings,mesh generation and solution,and post processing methods.The effect of buffer concentration and bias voltage on the DNA through nano holes is investigated in detail.The results are in agreement with the experimental results.A new simulation method is provided for the simulation of the magnetic bead binding DNA through nanopore.