Electrophoresis and DNA fingerprinting

Electrophoresis is a technique which uses an electric field to separate molecules, allowing for identification and characterization of the molecules. It is commonly used to separate nucleic acids and protein molecules of various sizes. To prepare the gel for electrophoresis the amount of agrose needed must be calculated. For a 0.8 percent gel 0.8 grams of agrose is necessary per 100 ml of buffer. The DNA fingerprinting experiment only called for 50ml of buffer, therefore only 0.4 grams of argose was needed. Once the buffer and argose were combined, the solution was microwaved until the argose had completely dissolved. While waiting for the solution to cool, the gel box was assembled by putting the gel tray into it and placing the gel comb at the end of the tray. As soon as the argose solution was cool enough to handle, our TA added ethium bromibe to our solution.

Ethium bromide is useful as a fluorescent tag (nucleic acid stain) for electrophoresis it binds to DNA and when exposed to UV light it glows bright orange. After the ethium bromide and the agrose gel were combined, the new mixture was poured into the gel tray and allowed to solidify. Once the gel became solid, it was carefully removed along with its tray from the gel box. Next, it was repositioned it so that the wells were on the negative side of the box. Since, DNA fragments are negative; this caused them to migrate away from the negative electrode. The electrophoresis device was then filled with 250ml of the 1X TBE buffer. The surface of the gel should be completely submerged. Finally, we removed the gel comb and added the sample.

After all the DNA samples were loaded, the cover was attached to the apparatus as well as the two wires from the power source. The process ran at 150V for roughly 40 minutes. During our experiment, the group on the other side placed their wells on the wrong side. This forced us to restart part way through our 40 minutes. Roughly after 40 minutes, the power source was turned off and the wires were unplugged. Carefully, the gel was removed and placed in a UV illuminator. Bands in the gel were easily visible. A picture of our results was taken and the gel discarded.

Figure : Gel Image of suspect and crime scene samples
Samples had been digested with restriction enzymes and run on a 0.8 percent gel.

Looking at Figure 1, we can see that row 3(crime scene two) matches up with row 7(suspect two). Suspect 1(rows 4 and 5) seem to match up with crime scene one (row 2). In this lab we used two different enzymes. This was important because restriction enzymes cut DNA into very precise recognition sequences. Every fragment of DNA has a specific number of nucleotides and base pairs. The reason we used “markers” in rows 1 and 8 was to determine the size of the DNA fragments produced with the enzymes we used. These “markers” acted as measuring tools. Notice in Figure 1 how the outside channels create a sort-of ruler. DNA fingerprinting, or electrophoresis, is used to separate out and therefore determine the different sizes of DNA fragments which are cut by restriction enzymes. Since restriction enzymes only cut at particular protein recognition sites, no two restriction enzymes will code for the exact the same site. This allows for a fingerprint-like uniqueness to every person’s DNA. By using this technique it is easy to find which DNA belongs to whom with a good degree of accuracy.

Judging by the data which was collected on the second crime scene, we can conclude that the second suspect was present at the scene. We can also see a correlation between the first suspect and the first crime scene. Although there is evidence of the suspects being present at the different crime scenes, we cannot automatically assume that they are the guilty party. All we can prove is that they were, in fact, at that specific location. Electrophoresis doesn’t have too many errors which can take place; however, human error is always a factor. If only one enzyme was used by mistake, distinguishing between the DNA would be much more difficult. This is due to the specific binding sites of each enzyme we used. They “snip” the DNA at different locations, allowing us to get a more accurate reading with the different strands of DNA. Cross contamination could be a possible error as well as poor injection of the sample into the gel.

One must have a sharp eye and steady hands to make it into the well without poking the gel. Another error which could have caused faulty results would be an incorrect measurement of agros in making the gel. If you had too much agrose in the gel your pore size would be smaller than it should have been. This would cause the DNA fragments to move much slower and not quite as far. DNA fingerprinting is a very useful tool. It is mainly used for and criminal investigation along with parental testing. DNA fingerprinting can also aid in detecting genetically inherited diseases. Some of the diseases it can help detect include hemophilia, cystic fibrosis, and Huntington’s disease. In many cases, if the disease is caught early enough there is a higher chance that it can be defeated or lessened. Several couples which are carries of certain genetic diseases seek out genetic counseling to help them understand the risk of having n affected child. DNA fingerprinting helps researchers as well by aiding them with patterns that each disease carries. These patterns make them easier to identify, which allows researchers to focus on specific areas of the DNA.

Works Cited
(2014, Oct. 14 ). In Electrophoresis. Retrieved Oct. 31, 2014, from http://en.wikipedia.org/wiki/Electrophoresis (Year, Month. Day ). In Lab 6B – DNA Fingerprinting. Retrieved Month. Day, Year, from http://www.biologyjunction.com/sample_6b_dna_lab_ap.htm Nakate, Shashank.. (2011, Jun. 20 ). In Electrophoresis Uses. Retrieved Oct. 31, 2014, from http://www.buzzle.com/articles/electrophoresis-uses.html