Biotechnology poster Gelelectrophoresis Gel electrophoresis is atechnique used to separate fragments of DNA, RNA or proteins by their length. Gelelectrophoresis is most commonly used for separating DNA fragments and willtherefore be discussed in most detail on this poster. In gel electrophoresisfor DNA sequences, restriction enzymes are used to cut the sequence at certaincombinations of nitrogenous bases. This gives rise to a number of DNA sequencesthat can then be profiled using a gel made of a polysaccharide called agarose. Theagarose gel contains small pores, which the fragments of DNA can travel throughand therefore be separated.
The DNA fragments are placed into the gel alongwith a ladder of fragments of DNA of known lengths that provide a control tothe test. These samples of DNA are movedthrough in the gel using an electric current. As DNA (and RNA) is negativelycharged, and sequences will have the same charge per mass, shorter sequenceswill move through the gel and towards the positive electrode at the bottom ofthe well more quickly than longer fragments. This creates columns of DNAfragments that can now be examined.
However, when separating proteins based ontheir lengths, an additional process is required as proteins are not negativelycharged. When using gel electrophoresis to separate proteins, researchers mustfirst mix the proteins with a detergent called sodium dodecyl sulfate. Thistreatment makes the proteins unfold into a linear shape and coats them with anegative charge. The bands found in the gel can bevisualized using a DNA-binding dye to stain the gel and UV light, which makesbands of DNA glow in the gel. Using the ladder of fragments of known base pairslengths makes it possible to identify the lengths of the bands that are beingtested. Gel electrophoresis has a widerange of applications in the real world. It is very commonly used in forensic scienceto match the DNA of suspects to that found at crime scenes.
Here, the DNA of asuspect will be used in the ladder of fragments that act as the control for theexperiment. If the DNA in the ladder matches that of the sample, then the twooriginate from the same person. Gel electrophoresis also plays and importantrole in research. For example, ordering DNA and RNA strands by length allowsresearchers to study them at a molecular level far more easily. Geneticists usethis technique to obtain a clearer picture of DNA and to prepare DNA for geneticengineering and cloning. Polymerase Chain Reaction In many cases, only a smallsample of DNA is available for testing.
When this sample isn’t sufficient forthese experiments to be performed, DNA needs to be multiplied. This is done bya process known as Polymerase Chain Reaction (PCR). PCR is a common laboratorytechnique used to make millions or even billions of copies of a particularregion of DNA. This allows a small sample of DNA to be analyzed throughsequencing or gel electrophoresis.As with DNA replication in anorganism, PCR requires a DNA enzyme that makes new strands of DNA from thetemplate strand. The DNA polymerase enzyme typically used in PCR is Taqpolymerase, named after the heat-tolerant bacterium from which it is obtained(Thermus aquaticus).
This enzyme is used as T. aquaticus lives in hot springsand hydrothermal vents, which means its DNA polymerase is very heat-stable andmost active at around 70°C.This is useful as high temperatures are constantly used in PCR to denature thetemplate DNA or separate its strands.