The Basics of DNA Purification

DNA purification is a vital component of many molecular assays which include PCR, qPCR, and DNA sequencing. It eliminates proteins that have been contaminated, salts, and other impurities that can hinder the downstream process. It also ensures the desired DNA is pure and available in a way that it can be used in future analyses. The quality of DNA is measured through spectrophotometry (the ratio of A260 to A280) and gel electrophoresis and many other methods.

In the initial step of a DNA purification procedure, the cellular structure will be disrupted with detergents or reagents such SDS in order to release DNA. To further purify DNA, reagents designed to denature proteins such as sodium dodecyl-sulfate or Ethylene Diamine Tetraacetic Acid (EDTA) can be added to denature them. The proteins are removed from the nucleic acids solution by centrifugation and washing. If RNA is present in the sample, a ribonuclease treatment could be added to further denature the RNA. The nucleic acids then are concentrated in ice-cold alcoholic to separate them from other contaminants.

Ethanol can be utilized as a solvent to remove salts and other contaminants from nucleic acid. A standard concentration of ethanol allows researchers to compare the results of different experiments, making it a suitable choice for workflows that require high-throughput. Other solvents, like chloroform and phenol, can be used but they are more hazardous and require additional steps to purchase science supplies prevent cross-contamination. Newer methods can make it easier to complete the DNA purification process using low-ionic-strength ethanol that has been shown to be as effective as conventional organic solvents in purifying DNA [2626. This is especially the case when paired with spin column-based extract kits.


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