Gel electrophoresis is a method used to separate pieces of
DNA in an
agarose gel. Molten
agarose is poured into a mould, and a comb is inserted in the liquid agar to form wells. Once the gel has cooled, a mixture of
DNA pieces, dye and buffer is put into each well, along with a reference lane. Before the
DNA can be added, however, it must be cut with
restriction enzymes.
The gel is then put into a buffer solution, in an apparatus that is connected to a power source. This buffer solution is conductive. This is required in order to get any movement of the
DNA. The electricity is turned on. Since
DNA is negatively charged, the molecules begin moving towards the anode, or positively charged end. Larger molecules will move more slowly through the gel, whereas smaller molecules can move much faster. Tracking dye is used to ensure that the
DNA does not run off the gel. The tracking dye is much, much smaller than any of the pieces of
DNA, and therefore will travel faster. Once the tracking dye nears the end of the gel, the power supply is turned off. This way, the
DNA will not fall out of the other side of the gel.
DNA is not visible on the gel, so a dye must be applied in order to be able to see the
DNA bands. The most common dye, and the one used in the Gene Technology workshop, is methylene blue. The gel is covered in methylene blue, which binds to the
DNA. The excess dye is washed off in warm water, and the
DNA bands are now clearly visible. Another option is to add ethidium bromide, a dangerous carcinogen, to the
DNA before running the gel. The ethidium bromide intercalates between bases, and glows under UV light. Once the gel has been run, it can be placed under a UV light to identify the bands of
DNA. This second method carries more safety hazards than the methylene blue method. The methylene blue method is usually sufficient for most gels, especially in a secondary school setting.
Applications of Gel Electrophoresis:
Gel electrophoresis is common in gene technology, and serves many different purposes. One of the most important is its use in forensic science, and solving crimes.
DNA fingerprinting is a method of comparing
DNA samples.
DNA can be extracted from blood, hair, or skin cells found at a crime scene.
DNA samples can be obtained from suspects. Both the original piece of
DNA and the suspects’
DNA can be run on a gel. On the gel, several loci are tested. It is rare that two unrelated persons will have an identical locus on a certain chromosome, however multiple loci are usually tested in order to ensure that the match is true. Only if all the loci are identical, is it considered a match.
Another application of
gel electrophoresis is in paternity testing. A child’s genetic markers will be a mixture of his or her mother and father’s. Any bands present in the child’s
DNA, but not in the mother’s
DNA, must belong to the father. By comparing the bands, the child’s father can be determined.
In genetic research, scientists use
gel electrophoresis when they want to know if a certain gene is in the
DNA they are looking at. There are a number of reasons why they would want to do this. The scientist could have inserted a foreign gene using a plasmid, and intend to study the effect of this gene on the target organism. However, the scientist must first ensure that the plasmid did indeed insert into the host organism’s
DNA. The scientist can extract the
DNA from the host, digest it using restriction enzymes and run it through a gel. Then, they can create a probe for a piece of their inserted gene. This is called a Southern blot, after its creator Edward Southern. The probe will attach to the target gene, and cause it to be seen. If the probe binds, the plasmid was successfully incorporated into the host
DNA.