Before making a PCR reaction or cloning test or DNA sequencing, it’s essential to have high-quality DNA which is free of contaminations such as protein, debris and RNA. Purifying DNA is also referred to as DNA Isolation and is a crucial step in molecular biology. This article will explain the fundamentals of DNA extraction and how to improve it for better results.
The first step in the process of purifying DNA is to prepare a solution which includes a mix of water and an alkaline buffer. This buffer makes the DNA soluble so that it can be easily separated from the other components of the sample. Once the DNA has been placed in a water and alkaline solution, it’s treated by chaotropic salts or detergents to dissolve cell membranes as well as nuclei and release DNA (cell lysis). RNase can be added to the sample to remove any DNA that is contaminating.
The DNA is then separated by organic solvents such as phenol or chloroform from other components of the cell, such as proteins and fats. After the DNA has been removed from proteins and lipids, it is able to be separated using ethanol or isopropyl alcohol (rubbing alcohol).
Spectrophotometry and Gel Electrophoresis can be used to determine the quality of DNA. A good quality DNA sample should have a ratio of absorbance of 260 nm up to 282 nm.
