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DNA Purification Methods

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15 Jul 2021

DNA Purification Methods

DNA (deoxyribonucleic acid) in every living cells are responsible for overall regulation of that cell (growth, survive and reproduce). On certain signals or stage of its life cycle the DNA gives messages to cell by transcription and translation. Each individual of certain species differs from each other on genetic basis and DNA is that genetic material in almost every living organism except of few viruses having RNA as genetic material.
DNA purification can be used for molecular analyses such as PCR, electrophoresis, sequencing genomes, fingerprinting, genetic engineering and cloning. It plays an important role in understanding the genetic cause of disease, detection of virus and bacteria. It is also essential for the development of diagnostics and drugs.
DNA extraction is the process of separating DNA from cells present in any biological sample. Mission is to separate DNA in the nucleus from other cellular components found in the cells. The extracted DNA should be of good quality and in efficient amount. Quality should be determined by purity of DNA without contamination such as RNA and proteins. Avoiding things like detergents, temperature, bad handling etc. which can denature DNA so should go through a certain protocol. DNA can be extracted by manual method as well as commercially available kits.

Step 1. Breaking cells open to release the DNA

The cells in a sample are separated from each other, such as by physical methods like vertexing, and put into a solution containing salt. The positively charged sodium ions in the salt protect the negatively charged phosphate groups that run along the backbone of the DNA.

A detergent is then added. The detergent breaks down the lipids in the cell membrane and nuclei. Hence DNA is released as when these membranes are disrupted.

Step 2. Separating DNA from proteins and other cellular debris

Often a protease is added to degrade DNA-associated proteins.

Step 3. Precipitating the DNA with an alcohol

Ice-cold ethanol or isopropanol added to the DNA sample. DNA is soluble in water but insoluble in salt and alcohol. So by stirring the alcohol layer with a sterile pipette, the precipitate is visible and can be taken out. If there is lots of DNA, you may see a white precipitate.

Step 4. Cleaning the DNA

The DNA sample can now be further purified. It is then resuspended in a slightly alkaline buffer.

Step 5. Confirming the presence and quality of the DNA Optical density readings taken by a spectrophotometer can be used to find out the concentration and purity of DNA in a sample. Also, gel electrophoresis can be used to show the presence of DNA in the sample.

54 thoughts on “DNA Purification Methods”

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