DNA-magnetic Particle Binding Analysis by Dynamic and Electrophoretic Light Scattering

Abstract

Isolation of DNA using magnetic particles is a field of high importance in biotechnology and molecular biology research. This protocol describes the evaluation of DNA-magnetic particles binding via dynamic light scattering (DLS) and electrophoretic light scattering (ELS). Analysis by DLS provides valuable information on the physicochemical properties of particles including particle size, polydispersity, and zeta potential. The latter describes the surface charge of the particle which plays major role in electrostatic binding of materials such as DNA. Here, a comparative analysis exploits three chemical modifications of nanoparticles and microparticles and their effects on DNA binding and elution. Chemical modifications by branched polyethylenimine, tetraethyl orthosilicate and (3-aminopropyl)triethoxysilane are investigated. Since DNA exhibits a negative charge, it is expected that zeta potential of particle surface will decrease upon binding of DNA. Forming of clusters should also affect particle size. In order to investigate the efficiency of these particles in isolation and elution of DNA, the particles are mixed with DNA in low pH (~6), high ionic strength and dehydration environment. Particles are washed on magnet and then DNA is eluted by Tris-HCl buffer (pH = 8). DNA copy number is estimated using quantitative polymerase chain reaction (PCR). Zeta potential, particle size, polydispersity and quantitative PCR data are evaluated and compared. DLS is an insightful and supporting method of analysis that adds a new perspective to the process of screening of particles for DNA isolation.

Contribution of magnetic particles in molecular diagnosis of human viruses

“…Simultaneous extraction of RNA or DNA from cancer cells consuming silica-coated MNPs was appropriate for future acts for instance, such as PCR as well as RT-PCR [ 84 ]. Immobilization of nucleic acid via SPRI (Solid‐phase reversible immobilization) onto MNPs proposes an elegant alternate for approaches based on centrifugation or else columns [ 85 ]. During drying, it is feasible to reversibly bind MNPs inside the sample with nucleic acids.…”

A novel method to optimise the utility of underused moulted plumulaceous feather samples for genetic analysis in bird conservation

“…Buffer AE contains the organic compound Tris [tris(hydroxymethyl) aminomethane, (HOCH 2 ) 3 CNH 2 )] and EDTA [Ethylenediaminetetraacetic acid (C 10 H 16 N 2 O 8 )] which functions to rehydrate the nucleic acids and release DNA from the silica membrane. This process is improved when the DNA and silica are exposed to higher temperatures (Bruns et al 2007;Zhou and Ling 2011;Haddad et al 2017). We used an elution volume of 100 µl in a two-step process, giving a final volume of 200 µl which was subsequently stored at − 20 °C.…”

A Robust, Safe, and Scalable Magnetic Nanoparticle Workflow for RNA Extraction of Pathogens from Clinical and Wastewater Samples

“…In the presence of a magnetic field nucleic acids are rapidly separated from most impurities, and the purified nucleic acids can be further released from the surface of MNPs by elution buffer with a different ionic strength. [ 28–31 ] SPRI allows fast and thorough washes to eliminate inhibitors of downstream molecular biology reactions and yields high quality RNA for PCR and high‐throughput sequencing. Because it requires no centrifugation and only low‐cost materials, the MNP‐based RNA extraction is inherently scalable and amenable to automation.…”