Necessary for a variety of applications in molecular biology, DNA extraction/isolation is a procedure whereby deoxyribonucleic acid (DNA) is exposed and removed* from a biological sample.
First accomplished almost 150 years ago, there are currently hundreds of protocols and kits optimised to perform the, now routine, procedure of DNA extraction, on a variety of samples. As different downstream applications have distinct requirements on DNA purity and integrity, choosing an appropriate DNA extraction method is of paramount importance for the success of the whole project.
In general, DNA extraction methods can be broken down into three stages:
- Tissue disruption and/or cell lysis
- Protein, RNA and other contaminant removal or destruction
- DNA recovery
1) Tissue Disruption and Cell lysis
Cell lysis is a critical stage in the DNA extraction protocol during which, cells are disrupted to release the nucleic acids.
While the DNA extraction procedure for many tissues and cells may begin with a simple lysis step, hard/tough tissues, such as bone and cartilage, may require additional milling or enzymatic pre-treatment, respectively. Alternatively, mixed samples may necessitate pre-processing, to separate the cells of interest prior to lysis.
Broadly speaking, cell lysis methods can be divided into three categories:
- Mechanical (e.g. grinding, ultrasonication, bead milling)
- Chemical (e.g. detergents, chaotropic agents)
- Enzymatic (e.g. proteinase K, lysozyme, lyticase)
To successfully break open the cells and nuclei, samples often require treatment using combinations of the above approaches.
Lysis reagents are often incorporated in commercially available DNA extraction kits. Alternatively, researchers may be required to lyse their cells prior to further processing, to remove unwanted materials and purify the DNA.
2) Protein, RNA and other contaminant removal or destruction
3) DNA recovery
These stages are highly variable depending on the method used. Please refer to “DNA extraction methods” for more information.
The suitability of a DNA extraction method is dependent on a number of factors including:
- Downstream application
- Source organism
- Sample material and quantity
- DNA type (e.g. plasmid or genomic DNA)
- DNA yield and average fragment size required
As such, no one method is superior; researchers are required to follow the procedure that best conforms to their laboratory set up, budget and the considerations of the aforementioned factors.
*→ The term “DNA extraction” may be used, exclusively, in reference to the removal of DNA from cells, or may also encompass DNA purification.
→The term “DNA extraction” is used interchangeably with “DNA isolation” and “DNA purification”. Alternatively, “DNA purification” may be used to describe methods of cleaning DNA, such as after enzymatic reactions, from gels etc.