The Basics of DNA Purification

Before performing the PCR reaction or cloning test or DNA sequencing, it is essential to have a high-quality DNA that is free of contaminations such as protein, debris and RNA. The process of purifying DNA is known as DNA isolation and is one of the most crucial steps in molecular biology. In this article, you will learn the basics of DNA purification and how to improve your DNA extraction processes for better results.

The initial step in the DNA purification process is to prepare a solution that contains a mixture of water and an alkaline buffer. This buffer makes the DNA soluble and so that it can easily be separated from other components of the sample. After the DNA has been placed in an alkaline-water solution, it’s treated with detergents and Chaotropics salts to break down cells’ membranes and nuclei. This lets the DNA out. RNase can be added to the sample in order to remove any contaminating DNA.

The DNA is then separated by organic solvents such phenol or chloroform from other components of the cell including fats and proteins. Once the DNA has been separated from proteins and lipids it can be extracted using ethanol, or isopropyl alcohol (rubbing alcohol).

Spectrophotometry and gel electrophoresis can be used to determine the quality of DNA. A good quality sample of DNA should have an absorbance ratio between the 260-nm range and 280-nm range. 1.8. A low ratio may indicate a problem with the protein binding processes or the transfer of salt from the bind or wash buffers.

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