DNA purification is a crucial process in a variety of molecular assays which include PCR or qPCR as well as DNA sequencing. It eliminates harmful proteins as well as salts and other impurities which hinder the downstream process. It also ensures that the desired DNA is in good condition and pure in order to be further analysed. The quality of DNA is determined through spectrophotometry (the ratio of A260 to A280) and gel electrophoresis and other methods.
In the initial step of a DNA purification procedure the cellular structure will be disrupted using detergents or reagents like SDS to release DNA. To further purify DNA, reagents that are protein denatured such as sodium dodecylsulfate as well as Ethylene diamine tetraacetic acids (EDTA) are added to denature bo finneman proteins. Then, they are removed from the nucleic acid solution using centrifugation and washing steps. If RNA is present in the sample, it can be further denatured by adding ribonuclease. The nucleic acids are concentrated in ice-cold alcohol to separate them from other contaminants.
Ethanol can be used as a solvent to remove salts or other contaminants from nucleic acid. Researchers can evaluate the results of different studies using an average ethanol concentration which is a good choice for workflows with high-throughput. Other solvents like chloroform and phenol, can be utilized, however they are more toxic and require additional steps to prevent cross-contamination. Newer techniques can facilitate the process of purifying DNA by using low-ionic-strength ethanol that has been shown to be equally effective as the conventional organic solvents when purifying DNA [2626. This is particularly relevant when used in conjunction with spin column extract kits.
