Isolation of DNA

Modern technology has led to a steady decrease in the amount of DNA needed for analysis.

Cell walls and membranes must be broken down to liberate the DNA from the cell.

Almost every molecular biologist has collected DNA from the organism they are studying. Initially, isolating DNA was a long and arduous process with large amounts of DNA collected. Advancing technology has resulted in the amount of DNA needed for either analysis or cloning of genes to steadily decrease. Nowadays, for example, enough DNA can be collected for genetic manipulations in laboratory mice or rats from a small piece of the tail. Human DNA may be analyzed using small blood samples or a few cells scraped from the inside of the cheek. The decrease in the amount of DNA required for analysis has allowed scientists to streamline the process so that DNA can be isolated in a few hours instead of a few days.

Extracting DNA from plants, animals, and bacteria, all require that the cellular contents be liberated into a solution. Since the bacteria are single cells and contain no bone, fat, gristle, etc., the DNA is relatively easy to extract. In contrast, samples from animals and plants must often be ground into tiny fragments before proceeding. Since plant cells have very rigid cell walls, the scientist must mechanically break the cells open in a blender, or add special degradative enzymes to digest the cell wall components. Similarly, to extract DNA from a mouse's tail, enzymes are added to degrade the connective tissue and disperse the cells. By far the easiest way to get DNA is to extract it from bacteria. A few drops of a bacterial culture will give plenty of DNA for most purposes. First, the bacterial cell wall is easily digested by lysozyme, an enzyme that degrades the peptidoglycan layer of the cell wall. A successive treatment with detergent dissolves the lipids of the cell membrane. Chelating agents, such as EDTA (ethylene diamine tetraacetate), are also used, especially with gram-negative bacteria, to remove the metal ions that bind components of the outer membrane together. In all these samples, the cellular contents, including the DNA, are then liberated into solution and are purified by a further series of steps.

Macromolecules such as DNA may be separated from smaller molecules by centrifugation.

Phenol dissolves proteins and is used to remove them from DNA.

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