Before making any PCR reaction or cloning experiment, or even DNA sequencing, it is essential to have a high-quality DNA source that is free of contaminants, such as debris, proteins and RNA. The process of purifying DNA is referred to as DNA isolation, and is one of the most critical steps in molecular biology. This article will teach you the basics of DNA extraction and how to optimize it to achieve better results.
The first step of the DNA purification process is to make a solution that consists of an emulsion of water and alkaline buffer. This buffer makes DNA soluble, which means it is easily separated from other components of the sample. After the DNA has been placed in an alkaline and water solution it’s cleaned with detergents as well as Chaotropics salts to break down the cell membranes and nuclei. This allows the DNA to be released. RNase may also be added to eliminate any contaminating RNA from the sample.
DNA is then separated from other cell components like proteins and lipids using organic solvents such as phenol and chloroform. Once the DNA is removed from the proteins or lipids it can be precipitated with alcohol or ruby alcohol.
The quality of the DNA can then be verified using spectrophotometry or gel electrophoresis. A high-quality sample of DNA should have an absorbance value between 250 nm and 280nm.