DNA purification is a crucial step in many molecular assays such as PCR, qPCR, and DNA sequencing. It removes contaminating proteins as well as salts and other impurities that can hinder the downstream process. It also ensures that the desired DNA is completely clean and present to be used in further studies. The quality of DNA is measured using spectrophotometry (the ratio of A260 to A280) or gel electrophoresis and other methods.
In the first step of a DNA purification process the cell structure will be disrupted using detergents or reagents like SDS in order to release DNA. To further purify DNA, reagents with protein denature such as sodium dodecyl sulfate or Ethylene diamine tetraacetic acid (EDTA) are added to break down the proteins. They then are removed from the nucleic acid solution using centrifugation and washing steps. If there is RNA in the sample, a ribonuclease treatment could be added to further denature the RNA. In the end, the nucleic acids is diluted using ice-cold alcohol to make it distinct from other contaminants.
Ethanol can be used as solvents to eliminate salts and other contaminants from nucleic acids. Researchers can compare results from different experiments by using an ethanol concentration that is standard, which is a good option for high-throughput workflows. Other solvents such as chloroform or phenol can be utilized, however they are more toxic and require additional steps to prevent cross-contamination with other proteins or cellular debris. The purification of DNA can be made simpler by using ethanol that has a low ionic strength. This has been shown to work as well as conventional organic solvents in cleaning DNA. This is especially the case when paired with a spin column extraction kit.