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Method Study on Magnetic Nanoparticles As Gene Engineering Carriers in Cell Transformation

Author: LiYao
Tutor: CuiHaiXin
School: Chinese Academy of Agricultural Sciences
Course: Biophysics
Keywords: Magnetic nanoparticles Carrier of genetic engineering PK-15 cells Arabidopsis protoplasts
CLC: Q78
Type: Master's thesis
Year: 2010
Downloads: 65
Quote: 0
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Abstract


Using magnetic nanoparticles as a carrier of genetic engineering of transgenic plant and animal cells, a polyethylene imine (PEI) is selected magnetic nanoparticles, aminosilane magnetic nanoparticles and chitosan magnetic nanoparticles three magnetic nanoparticles as gene engineering carrier, on the characteristics of their morphology, conversion effects in animal and plant cells and its influencing factors were discussed. The current initial results obtained are as follows: (1) by scanning electron microscopy, the morphology of the three magnetic nanoparticles characterization, three types of magnetic nanoparticles showed a spherical structure, PEI magnetic nano-particle size of 100-200nm, well dispersed, the aminosilane magnetic nanoparticles and chitosan magnetic nano particle size is about 50nm serious agglomeration. Analysis by gel retardation assay and the binding rate, that three magnetic nanoparticles can bind DNA, wherein the PEI magnetic nanoparticles DNA binding ability, the the aminosilane magnetic nanoparticles and chitosan magnetic nanoparticles DNA binding capacity is relatively weak. Further PEI magnetic nanoparticles with DNA binding was analyzed by atomic force microscopy and circular dichroism, DNA and PEI magnetic nanoparticles can be combined to form a tight composite structure. The complex formed by the magnetic nanoparticles with DNA by DNase I digestion, and found PEI magnetic nano-particles having the ability to protect the DNA, while the weaker aminosilane magnetic nanoparticles with chitosan magnetic nanoparticle's ability to protect DNA. (2) MTT law of magnetic nanoparticles on the toxicity of the porcine kidney cells (PK-15) cells, three kinds of magnetic nanoparticles on PK-15 cell toxicity is low, wherein of PEI magnetic nanoparticles toxicity relative maximum, aminosilane magnetic nanoparticles and chitosan magnetic nanoparticles toxicity is relatively small, cell survival rates were 68%, 83% and 87%. Magnetic nanoparticles transfected PK-15 The experimental results show that PEI magnetic nano-particles has a high conversion efficiency, when the PEI magnetic nano-particles with the DNA mass ratio of 1:1, the highest transfection efficiency to 25%, relative to the The higher the conversion efficiency of the liposome a positive control (10%), fluorescence stronger. Chitosan magnetic nanoparticles conversion efficiency is relatively low, at about 3%, aminosilane magnetic nanoparticles transfected with no expression. The experiments show that PEI magnetic nanoparticles for the PK-15 cells so that a gene vector. (3) For the first time using magnetic nanoparticles as gene vectors carrying DNA was transformed the Arabidopsis protoplasts obtained successfully expressed. Experiments found that the toxic effect of PEI magnetic nanoparticles on protoplast larger, when the amount of the magnetic nanoparticles in the 10μg, protoplast intact rate at about 10%, the aminosilane magnetic nanoparticles and chitosan magnetic nanoparticles is less toxic protoplast intact rate above 74% and 81%, respectively. Magnetic nanoparticles into Arabidopsis protoplasts experimental results show, PEI magnetic nanoparticles has certain transformation of Arabidopsis protoplasts, but the conversion efficiency is lower when the PEI magnetic nanoparticles with DNA mass ratio of 1:2 the highest conversion efficiency of about 11%, far below the 65% of the conversion efficiency of PEG positive control. Aminosilane magnetic nanoparticles and chitosan magnetic nanoparticles into Arabidopsis thaliana The protoplast no GFP expression.

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CLC: > Biological Sciences > Molecular Biology > Genetic engineering (genetic engineering)
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