Evaluation of Nucleic Acid Extraction and Purification Methods
PC (P is the abbreviation of English phenol, C is the abbreviation of English chloroform) extraction/alcohol precipitation method is a method that will never be outdated. Stable, reliable, economical and convenient. PC extraction can completely remove proteins, and alcohol precipitation can remove salts. For ordinary clean samples (impurities are proteins), this method can completely obtain high-quality nucleic acids. Although each PC extraction will lose a part of the nucleic acid (because it is impossible to remove all of the water phase), and the low concentration of nucleic acid alcohol precipitation efficiency is low, but these problems can be solved by operating adjustments or reduce the impact.
The biggest problem
The biggest problem of this method is that it is not suitable for large-scale extraction. PC extraction is a very effective means to remove protein. Phenol can denature the protein, and the denatured protein is separated from the water phase, in the phenol or between the phenol/water phase. The key to PC extraction is to mix thoroughly and to use enough. Thorough mixing can ensure sufficient contact between phenol and protein and completely denature the protein. Many people always worry about whether the violent mixing will cause damage to nucleic acids, especially genomic DNA. In fact, there is no need to be so careful. 
Vigorous mixing operation will partially disrupt the genomic DNA of large molecules, but the damage will not be so strong that the DNA becomes a small fragment within 10kb. After shaking and mixing vigorously, most of the genomic DNA fragments will be larger than 20kb. This size, except for some special requirements, is completely suitable for PCR and restriction digestion. If the required fragments are very large, such as to construct a library, you cannot use a violent mixing method, but only gently inverted and mixed back and forth. 
The key at this time
The key at this time is: the ratio of the lysis solution should be large enough to make the system not too viscous. The amount should be sufficient because phenol has a certain degree of saturation to remove protein. If the saturation is exceeded, the protein in the lysis system will not be removed at one time, and it must be extracted multiple times before it can be completely removed. In addition, the disadvantage of the system being too viscous is that the protein is difficult to completely remove, and the genomic DNA will be broken more severely, so pay attention to the ratio of the lysate to the sample. The 4C centrifugal operation is conducive to more thorough removal of protein. Another use of PC extraction is to use acidic phenol to partially remove DNA, and to obtain RNA with minimal DNA residue during RNA extraction. However, one thing to remind is that some plant samples cannot be extracted with PC before some impurities are removed, otherwise the nucleic acid will be degraded.
The high-salt precipitation protein/alcohol precipitation method
The high-salt precipitation protein/alcohol precipitation method is also a very good method. Compared with the PC extraction method, this method almost overcomes all the shortcomings of PC extraction, except that the purity stability may be lower. The benefit of faster and easier removal of protein is that it can be used for large-scale extraction, but the disadvantage is that the purity (protein residue) is not stable enough. The precipitation efficiency of protein is better at 4C.
The media purification method is a method that has received more and more attention. Its biggest feature is that it is very suitable for large-scale nucleic acid extraction, and because it is not affected by human factors, the stability of the purity is high. Its Achilles’ heel is the excessive amount of sample. The medium can be divided into two categories, one is column type, that is, the medium is pre-filled in the column below it; the other type is granular. The purification operation of the granular medium is not much different from the classic alcohol precipitation. It is through the process of adding liquid and pouring liquid several times. After drying, the purified nucleic acid can be obtained by dissolving it.
Although the operation of column purification also has the process of adding liquid and pouring liquid, because the added liquid will enter another centrifuge tube after centrifugation, and it is completely separated from the column containing nucleic acid, so the washing is more thorough and the operation is more labor-saving. However, the cost of the media purification method is the highest.
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