Tris (trimethylolaminomethane) is a common biological buffer, which can be used in the whole DNA extraction process. DNA is sensitive to pH during extraction from multiple sources. Tris can be used to maintain a stable pH during cell lysis, removal of unwanted cell components and precipitation. In addition, it also plays a particularly important role in cell lysis.
Tris protects DNA from pH changes
Tris can maintain a stable pH value, with pKa of 8.1 and effective buffer range of 7 to 9. Due to its neutral range, Tris is a common buffer in biological laboratories. When cells divide, their DNA and contents overflow into the buffer. RNase A (destruction of RNA), protease (destruction of protein) and SDS (sodium dodecyl sulfate, dissolution of membrane fragments) as well as the soup of fragmented RNA and protein have a great impact on the pH value of the solution. Because DNA is sensitive to pH, it is important for tris to buffer and stabilize pH.
Tris can promote cell lysis
Lysis or cell lysis is the first step of DNA extraction. This can be done by a buffer containing Tris and EDTA (ethylenediamine tetraacetic acid). EDTA binds divalent cations such as calcium and magnesium. Because these ions help to maintain the integrity of the cell membrane, eliminating them with EDTA will destroy the stability of the cell membrane. Tris is the main buffer component. Its main function is to maintain the pH value of the buffer at a stable value (usually 8.0). In addition, Tris may interact with LPS (lipopolysaccharide) in the membrane, which further destroys the stability of the membrane.
DNA must be in Tris buffer to be used
In the final stage of DNA extraction, DNA itself is extracted from the solution. At this time, DNA can be dissolved in buffer. In order to extract from the solution, DNA can be made insoluble by adding ethanol or isopropanol (isopropanol). After this operation, the DNA will appear as a white linear substance in the solution. Although DNA can be isolated from the remaining cellular components in this way, it is "unavailable" when DNA is insoluble. After separation, the alcohol is removed, and the DNA must be put back into biological buffer (such as Tris) before it can be used.
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