A high-quality DNA sample is required for a variety of downstream applications like PCR and sequence. DNA purification is the process of removing contaminants such as proteins, and other cellular elements, from a sample to produce a clean nucleic acid solution ready for use. There are a myriad of DNA purification methods to choose from each with distinct advantages and disadvantages, depending on the starting material and the intended application.
The first step in DNA purification is to remove proteins from the sample by using a proteinase (protein enzyme) or mechanical disruption. After eliminating the cellular debris, the DNA is precipitated in the presence of ethanol, which results in a white, stringy precipitate. The precipitated DNA can then be resuspended using water or a sterilized solution. The concentration of DNA can be determined using the spectrophotometry method based on the peak of nucleic acid absorbance at 260nm.
Salting out is yet another popular method used to purify DNA. A cellulose column is used for this procedure to capture and hold DNA. The blog cellulose matrix has to be pretreated with detergents to get rid of contaminants. A wash buffer is then applied to eliminate the salts. DNA binds to matrix in low salt conditions and contaminating proteins and RNA can be cleaned by using higher salt solutions. The eluted RNA and DNA are then recovered by precipitation with ethanol.
Anion exchange is also a popular method of purifying DNA. This method employs an agent that attracts positively charged molecules of DNA, while an agent that neutralizes negatively charged molecules of DNA is used to wash out the column. Once the DNA is eluted it can be concentrated by centrifugation. Then, the DNA can be removed by washing it off with 70 percent ethanol at ice-cold temperature.