Before making the PCR reaction or cloning experiment, or even DNA sequencing it is critical to have a high-quality DNA source that is free of contaminants such as protein, debris 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 improve it for better results.
The initial step in the process of purifying DNA is to prepare a solution that contains a mixture of water and an alkaline buffer. This buffer makes DNA soluble, and it is able to be separated from other components of the sample. After the DNA is placed in an alkaline-water solution, it is then treated with detergents and chaotropics salts to break up cell membranes as well as nuclei. This allows the DNA to be released. RNase may also be added to remove any contaminating RNA from the sample.
The DNA is then separated by organic solvents, such as chloroform and phenol from other components of the cell like fats and proteins. Once the DNA is removed from the proteins or lipids and lipids, it is then precipitated using ethanol or ruby alcohol.
The quality of the DNA can be determined by spectrophotometry or gel electrophoresis. A good quality DNA sample should have an absorbance ratio between 250 nm and 280nm. 1.8. A low ratio may indicate problems with the protein binding processes, or a salt carryover from wash or binding buffers.
Add a Comment