Preparation Of Magnetic Fe₃O₄/SiO₂ Composite Nanoparticles And Study Of Its Application In DNA Purification

In recent years, along with the development of nano materials science, a new domain has been opend about biochemistry separation technology. Magnetic composite nanoparticles were used to separate nucleic acid, protein et al. This kind of technology have many advantages and bright future that traditional technology does not have.The main content of this article were the preparation and characterization of magnetic Fe₃O₄/SiO₂ composite nanoparticles and its application in DNA purification.The results are:1. Magnetic Fe₃O₄/SiO₂ composite nanoparticles were prepared by the improved chemistry coprecipitation method. The average diameter of the Fe₃O₄ nanoparticles wasabout 6nm～12nm as confirmed by TEM. The dispersivity of Fe₃O₄ nanoparticles were very good. Superparamagnetic particles's aturation magnetization was 62.6emu/g, XRD patterns of the magnetic nanoparticles showed that the samples exhibit identical cubic inverse spinel structures. We assigned the absorption band at 578cm^-1and 3421cm^-1 tally with the standard Fe₃O₄ characteristic absorption peaks.2. No.1 and No.2 magnetic Fe₃O₄/SiO₂ composite nanoparticles were prepared by sol-gel method for one time( No. 1) and two times( No. 2).The material were TEOS, ethanol and ammonia . The average diameter of the No. 1 nanoparticles were about 20nm～30nm and No.2 are 30nm～50nm as confirmed by TEM. The dispersivity of both nanoparticles were very good. Superparamagnetic particles' saturation magnetization were 55.2emu/g and 51.8emu/g. XRD patterns of the magnetic nanoparticles showed that the samples exhibit identical cubic inverse spinl structures. So the low angle peak present in nano composite suggested the silica was without much indication of ordering.We assigned the absorption band at 1097cm^-1 and 950 cm^-1 tally with the standard SiO₂ characteristic absorption peaks.3. There exists difference of No.1 and No.2 magnetic nanoparticles in ability of combining with DNA in binding buffer of best matching ratio (20% PEG and 4 mol/L NaCl blend liquid). No.1 and No.2 magnetic nanoparticles were used to recover 500ng standard DNA sample.Ratio of DNA recovery is 78.2% which was a little higher than 71.6% of No.1. Therefore No.2 magnetic nanoparticles were used in following tests .Amount of captured DNA of 100μg No.2 magnetic nanoparticles was 519ng. Adsorbed DNA was eluted with the TE solution, its elution ratio was reaching as high as 95%-97%.4. No. 2 magnetic Fe₃O₄/SiO₂ composite nanoparticles were used to extract plasmid DNA:classical alkaline lysis method was improved. PEG and guanidine hydrochloride were used as binding buffer respectively. And magnetic method were used after Lysozyme lysis. Three magnetic methods were contrasted with KIT method. 10.2μg,8.5μg,8.2μg plasmid DNA were extracted by three magnetic methods and 9.5μg by KIT method. The plasmid DNA had superhelix structure and were complete mostly. OD260/OD280 ratio was 1.8±0.1. The plasmid DNA could be used for PCR and other molecular biology operation.5. No.2 magnetic Fe₃O₄/SiO₂ composite nanoparticles were used to extract animal blood genomic DNA: magnetic method traditional method and KIT method were compared in purity, quantity, speed and price. 8.1μg, 10.1μg and 8.3μg genomic DNA could be extracted from 200μL rabbits'blood. 5.0μg, 7.6μg and 5.6μg genomic DNA could be extracted from 200μL chickens' blood. 20 samples could be extracted in 50min,50min and 12h. Agarose eletrophoresis and PCR amplification showed that all the three methods could be used in the following molecular biology application. Every sample cost 0.4 Yuan, 40 Yuan and 4 Yuan with these three methods.Chickens'genomic DNA extracion didn't need centrifugation.6. No.2 magnetic Fe₃O₄/SiO₂ composite nanoparticles were used to extract DNA from agarose gel. When NaI solution concentration is 6 mol/L, pH=5.0,the Quantity One software indicated that the recovery efficiency was 60% between 750bp and 2000bp and 20%～60% smaller than 750bp.