Nowadays, there is a wide selection of commercially available kits, which operate using a combination of solvents and solid-phase support columns/beads to extract DNA from a variety of samples.
A rapid and widely employed technique, this method enables the extraction of high-molecular weight DNA via solid-phase anion-exchange chromatography.
This technology exploits the electrostatic interaction between the negatively charged phosphates of the DNA backbone and positively charged molecules on the surface of a substrate (e.g. an anion exchange resin in a spin column). Under low salt conditions, DNA binds specifically to the substrate while contaminants (such as RNA and proteins), are subsequently removed during wash steps using low or medium salt buffer. The remaining DNA is then eluted from the column using a high-salt buffer, and recovered via alcohol precipitation.
Anion-exchange technology is commonly employed in plasmid extraction kits. This technology delivers ultra-pure DNA, which can be used in a number of sensitive applications, and does not require lengthy procedures or toxic substances. Limitations to this method include the need for a centrifuge and an alcohol precipitation step.
In a simple bind-wash-elute procedure, DNA extraction with silica matrices is a popular method for the isolation of high-quality DNA via adsorption chromatography.
Commonly sold in a comprehensive kit with lysis and other buffers, silica technology can be incorporated in columns, microchips and beads. Using this method, DNA is adsorbed onto silica beads, particles or membranes, at a defined pH, in high concentrations of chaotrophic salts. The binding of DNA to silica matrices is strong, but reversible; it occurs as a result of the interaction between the negatively charged DNA backbone and the positively charged silica particles. In optimal conditions, the DNA remains bound to the silica matrix, and contaminants are washed away. Following washing, a low salt buffer is used to disrupt the DNA-silica binding, and the DNA is eluted from the membrane and collected, ready for use (i.e. without an alcohol precipitation step).
Silica based technologies provide a simple and rapid solution to DNA extraction, which can also be automated. However, this method is not suitable for all downstream applications; e.g. the DNA isolated is not “transfection-grade”.
The newest of the solid-phase DNA extraction methods, magnetic separation is a highly simple and rapid technique, which can be automated for high-throughput applications.
This technique utilises magnetic beads/particles, coated with a surface that interacts specifically and reversibly with DNA (e.g. silica), or DNA-binding antibodies. The procedure involves incubating a buffered cell lysate with magnetic beads, prior to applying a magnetic field to separate the beads (and bound DNA) from the unwanted cellular components. The unwanted materials are subsequently removed, the beads are washed and the DNA is eluted - ready for use.
While the aforementioned solid-phase technologies also deliver fast and simple DNA extraction procedures, magnetic separation is advantageous as it does not require any centrifugation steps. However, this method requires the use of specialised automated instruments. Furthermore, the reagents present in the final elution buffer may not be compatible with sensitive downstream applications, such as transfection.
DNA extraction has become an essential and routine procedure, necessary for a number of life sciences research, medical diagnostic, and forensic applications worldwide. As is evident, no one method is appropriate for all applications. Consequently, new methods and technologies; aimed at increasing purity and speeding up the DNA extraction process, are constantly emerging.
Birnboim, H.C. and Doly, J. (1979) A rapid alkaline lysis procedure for screening recombinant plasmid DNA. Nucleic Acids Research. 7: 1513.
Birnboim, H.C. (1983) A rapid alkaline extraction method for the isolation of plasmid DNA. Methods Enzymol. 100: 243.
Tan, S.C. and Yiap, B.C. (2009) DNA, RNA, and Protein Extraction: The Past and The Present. Journal of Biomedicine and Biotechnology. 2009: 1.
Uhlen, M. (1989) Magnetic Separation of DNA. Nature.340: 733-734
Hawkins, A. (1998) DNA purification and isolation using magnetic particles. http://www.google.com/patents/US5705628. Google Patents.