Genetics Laboratory Techniques
In order to analyze DNA, it is necessary to isolate it from its source. Often times, this source will be whole cells from organisms, which in addition to DNA, will also contain lipids, proteins, signaling molecules, and other types of compounds. Pure DNA is obtained through extraction of it from the rest of the cell, separating it from other things in the cell and allowing for analysis of the pure DNA. Traditionally, this has been done using a process known as Phenol extraction, which lyses the cell and precipitates DNA and nothing else in the cell, allowing us to obtain DNA through a series of mixing and fractioning. In recent years, however, companies such as Qiagen have created kits that allow for simple extraction of DNA without the long tedious process normally required by Phenol extraction. This allows geneticists to easily and quickly obtain pure DNA for subsequent analysis.
These subsequent steps for analysis of the pure DNA, however, requires the DNA to be in large amounts. While organisms typically have many cells from which to extract DNA, the extraction process is not fully accurate, and some cells have different sets of DNA due to processes such as mitotic recombination. Furthermore, our current techniques of analysis allow us to analyze specified lengths of DNA (<1000 base pairs) at any given location. Thus, it would be convenient to have a large, pure pool of a specific location in the genome of interest. This is obtained through a process known as Polymerase Chain Reaction(PCR). PCR involves the amplification of a portion of the DNA (<1000 base pairs) on an exponential scale. It involves repeated processes of melting the double stranded helix, annealing of primers, synthesizing the complement strand using polymerase, and yielding two strands of DNA identical to the first. Thus, with each cycle of this, the number of strands of DNA available doubles. The process is depicted in simplicity in this diagram:
Once a decent amount of DNA has been produced, the process of analysis begins. One of the most important techniques involved in analysis of DNA is the ability to separate DNA by size on a tray of agarose or polyacrylamide gel. Because DNA have a negative charge to their double helix backbone, they have a tendency to migrate towards the positive electrode when an electric current is applied. The gel allows geneticists to separate DNA by size when an electric field is applied. Longer pieces of DNA travel faster and will migrate through the gel farther than larger pieces of DNA. This provides the basis for techniques such as electrophoresis, and DNA sequencing. Electrophoresis allows separation of DNA by size, producing bands such as these on a gel plate:
More importantly, however, the process of separating DNA using gels have become the basis for the process of DNA sequencing, which is often the purpose of large-scale projects such as the Human Genome Project.
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