| With the development of biological science,people know that gene is genetic code of humanity and protein is executor of all biological activities.Reading all the gene information means we can understand the essence of our life.Meanwhile,mastering information of protein in our body let us know the physical status.Nowadays,nanopore is the most ideal tool to detect both of them.The third generation of DNA sequencing is based on this technique and has been considered as the vital means for "Precision Medicine Initiative" in the future.In this thesis,a series of experiments are conducted to confirm the feasibility of detection of biological molecules by solid-state nanopore,and research the principle that DNA and protein translocate through nanopore.The main results are presented as follow:1)The double strand ?-DNA(called DNA for short)passing through pore is studied in two conditions:one is the same concentration but different sorts of electrolytes,the other is the different concentration of LiCl.The spatial configurations of DNA strand is analyzed through nanopore.The results demonstrate that the dwell time of DNA in the pore is relative to ionic binding strength and quantity.The net charge will be less with the increasing of ions binding on surface of DNA,and the dwell time will be longer.Besides,DNA strand can translocate through in straightened and folded configuration.2)The behavior of BSA translocation through nanopore is researched to investigate the general theory of protein passing through pore.Under the different applied electrical field,the mechanism of protein unfolding is studied.Besides,the interaction between wall of pore and protein is discussed and the frequency of translocation events is analyzed.It is found that BSA will transform from folded state to partially unfolded state then to completely unfolded state.The transformation from one state to another one is an abrupt rather than a gradual process when the electrical field reaches a certain value.Moreover,in KCl solution,it is more easily for BSA to passing through pore.However,BSA also more likely stick into the inner wall of nanopore.The condition is reverse in LiCl.3)The mixture of BSA and DNA is detected,and then we contrast the results with the data of BSA and DNA separately.According to the trace of ionic current,it's more similar to the phenomenon in DNA experiment.While some long time events which usually occur in BSA experiment are discovered,but the quantity is too small.The long time events in BSA results can be distinguished with the events in DNA results.4)The ability of detection complex biological molecules is investigated using solid-state nanopore,such as IgG,IgA and IgM.Results indicate that those molecules are hard to get into nanopore.But,they will wander around the entrance of pore and produce collision signals.Moreover,the larger the molecular weight of them is,the higher the collision signals take up the ratio of all events.5)To enhance the ability of discrimination biological molecules by nanopore and optimize the experimental results,new method is explored to improve the ratio of signal-to-noise.It is found that the salt gradient can enlarge the amplitude of blockage current when charged molecules translocate through pore from high concentration side to low concentration side.Besides,the amplitude of blockage current becomes larger with the increase of salt gradients across the nanopore.However,it has a little effect on the dwell time.6)The finite element Multiphysics software(COMSOL)is utilized to simulate the fluid condition in nano-channel under asymmetric salt conditions.According to simulation results,it is deduced that the amplification effect of blockage current is relative to three factors:the surface charge of biological molecules,the ion diffusion under salt gradient and the electrical double layer.Lastly,different quantity of charge in molecules is set to confirm the simulation results.And the last result proves our deduction. |
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