Before doing the PCR reaction or cloning test or DNA sequencing, it’s essential to have a high-quality DNA that is free of contaminations such as debris, proteins and RNA. Purifying DNA is also referred to as DNA Isolation and is a vital step in molecular biology. This article will guide you through the fundamentals of DNA extraction and how to optimize it for better results.
The initial step of the DNA purification process is to prepare a solution containing an amalgamation of water and alkaline buffer. This buffer makes the DNA soluble, so that it can easily be separated from the other components of the sample. After the DNA has been placed in an alkaline-water solution, it is then treated with detergents and Chaotropics salts to break down cell membranes and nuclei. This lets the DNA out. RNase may also be added to eliminate any contaminating RNA from the sample.
DNA is then separated from other cellular components like proteins and lipids with the help of organic solvents such as phenol and chloroform. Once the DNA is removed from lipids or proteins it is then precipitated using alcohol or rubbing alcohol.
The purity of the DNA sample can be determined by spectrophotometry or gel electrophoresis. A good quality sample of DNA should have an absorbance ratio between the 260-nm range and 280-nm range.