What is the function of Tris buffer in DNA extraction?
Tris (Tris(hydroxymethylaminomethane) is a common biological buffer that can be used for the entire DNA extraction process. DNA is sensitive to pH during extraction from multiple sources. In the process of cell lysis, removal of unwanted cell components and precipitation, tris can be used to maintain a stable pH. In addition, it also plays a particularly important role in cell lysis.
Tris protects DNA from changes in pH
TRIS can maintain a stable pH with a pKa of 8.1 and an effective buffer range of 7 to 9. Due to its neutral range, Tris is a commonly used buffer in biological laboratories. When cells divide, their DNA and contents spill into the buffer. RNase A (destroy RNA), protease (destroy protein) and SDS (sodium dodecyl sulfate, dissolving membrane fragments). This cell content and the soup of fragmented RNA and protein have a great effect on the pH of the solution. influences. Because DNA is sensitive to pH, it is important that Tris buffer and stabilize the pH.
TRIS can promote cell lysis
Lysis or cell lysis is the first step in DNA extraction. This can be done with a buffer containing tris and EDTA (ethylenediaminetetraacetic acid). EDTA binds divalent cations (such as calcium and magnesium). Since these ions help 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, and its main function is to maintain the pH of the buffer at a stable value (usually 8.0). In addition, Tris may interact with LPS (lipopolysaccharide) in the membrane, thereby further destabilizing the membrane.
DNA must be in tris buffer before it can be used
In the final stage of DNA extraction, the DNA itself is extracted from the solution. At this time, the DNA can be dissolved in the buffer. In order to extract from the solution, the DNA can be made insoluble by adding ethanol or isopropanol (isopropanol). After this operation is completed, the DNA will appear as a white thread-like substance in the solution. Although DNA can be separated from the remaining cellular components in this way, it is "unusable" when the DNA is insoluble. After separation, the alcohol is removed, and the DNA must be returned to the biological buffer (such as tris) before it can be used.
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