Gold Magnetic Particles As The Carrier Of Whole Blood Genomic Dna Purification Methods

Magnetic composite particles are superparamagnetic colloid composed of macromolecule/inorganic materials and magnetic nanoparticles(such as iron oxide,iron, cobalt,nickel and other metal).With the superparamagnetic property of magnetic core and the nature biomolecule affinity of the gold surface,Fe₃O₄/Au magnetic composite particles (named as GoldMag(?) particles) became one of the research hotspots.Because of small size, large specific surface area,superparamagnetic and easy biomolecule affinity,GoldMag(?) particles has been widely used in immunology detection,enzyme immobilization,protein separation and other biological fields.In this study a method of genomic DNA purification with GoldMag(?) particles as a carrier was established.The purification steps of lysis of whole blood,amount of GoldMag(?) particles added, DNA binding on magnetic particle surface,washing and eluting,were optimized.Using human blood,rat blood and ?blood as samples,the genomic DNA purification protocol from the whole blood of mammalian was established,the method was evalued by Uv-vis aborption, electrophoretic ananysis,PCR etc.;The genomic DNA purification protocol from birds blood, was also set up purified from chicken blood by adjusting the mammalian protocol.The results showed that 2～5μg genomic DNA was yield from 100μL human whole blood;20μg genomic DNA was yield from 5μL chicken whole blood.The A₂₆₀/A₂₈₀ ratios of obtained genomic DNA were in the range of 1.70～1.90.The CV(coefficient of variance) in a batch is lower than 7%,The CV of between different batchs is lower than 20%.The purification can be performed in 50 min and easy to operate without toxic reagents such as phenol or chloroform,showing the same high quality as similar method.A background absorbance at 260 nm was detected with ultrapure water as a control sample which process using the purification steps.The result showed that the background was due to the reaction between Fe₃O₄(the core of GoldMag(?) particles) and EDTA(in elution buffer) to generate a water soluble chelate complex,which could be accelerate by high temperature.The comparation of GoldMag(?) particles with Fe₃O₄ magnetic particles showed that the background of Fe₃O₄ was 10 times higher than that of GoldMag(?) particles,indicating the Au nanaparticles assembled on GoldMag(?) particle surface greatly prevented the ironic chelation with EDTA.However,Seleting Tris-HCl as elution buffer or using TE buffer at room temperation could eliminate the background.Fortunately,the background has no influence on the downstream application of purified genomic DNA.