Gel Electrophoresis Lab

William Poteat Biology P.4 Lab Write-Up 5/19/14 Gel Electrophoresis and DNA Fingerprinting Lab Write-Up Purpose: The purpose of this lab was to investigate the process used for DNA fingerprinting by means of Electrophoresis methods. We examined and learned/applied how the process was done, and successfully examined and analyzed the travel patters of the “DNA samples” in electrophoresis gel. Materials and Methods: Our materials included, but are not limited to: an electrophoresis gel, a comb, an electrophoresis tank, with both +/- sides, samples Bromephenol Blue, Methyl Orange, Ponceau G, Xylene Cyanol, Pyronin Y, Unknown 1, 2, and 3, a battery to power the tank, sample injectors, data sheets, spare sample collection tubes, spare wires, a ruler, and an extra plate on which to put the completed electrophoresis gel. In order to carry out our experiment, we first made the electrophoresis gel, by using a comb to create sample injection areas. We then placed the gel into an electrophoresis tank. We inserted all of the samples into the gel using the sample injectors and spare sample collection tubes. When this was completed, we plugged the electrophoresis tank into the battery using spare wires. We waited approximately 10-20 minutes then removed our gel. We placed it onto out extra plate, measured the sample travel distance through the gel with our rulers and collected our information onto our data sheets for analyzation. Introduction: 1.) Gel Electrophoresis is done by inserting samples into an electrophoresis gel then applying electrical current in order to move the samples in certain charged directions based on their size. It separates the DNA into a pattern of bands because it sorts the larger pieces from the smaller or other sized pieces. The travel distance is based on charge and size of the DNA pieces/ molecules. 2.) The DNA samples are processed by being cut by restriction enzymes. Essentially, these enzymes locate specific points on the DNA bands and cut them in order to separate the useful pieces from the useless pieces. 3.) The names of the dyes are shown above. We were able to figure out the composition of each unknown dye by looking at the distance traveled compared to the other dyes, and calculating which mixture would effectively give us these unknowns. The unknowns were composed of: Unknown 1- Xylene Cyanol and Bromephenol Blue Unknown 2- Xylene Cyanol and Ponceau G Unknown 3- Methyl Orange and Pyronin Y

DATA: Bromphenol Blue: Bands; 1, Positive movement, 2cm Methyl Orange: Bands; 1, Positive movement, 1.25 cm Ponceau G: Bands; 1, Positive movement, 4cm Xylene Cyanol: Bands; 1, Negative movement, 2cm Unknown 1: Bands; 2, positive movement, .75, 2.5 cm, (Xylene Cyanol + Bromphenol Blue) Unknown 2: Bands; 2, Positive movement, 3, 3.5 cm, (Xylene Cyanol + Ponceau G) Unknown 3: Bands; 2, Pos. + Neg. movt., 1, 2cm, (Methyl Orange + Pyronin Y) Analysis and Conclusions: After completing this lab, we can conclude that dyes travel in opposite directions based on their charge. For example, a positively charged dye will travel negatively and vice versa because opposite charges attract. The graph for some reason shows no correlation between dye size and distance traveled which is surprising because it is not what one would expect. The composition of the unknowns is said above. I am not confident the experiment correctly identified unknowns because we were unable to find exact matches and there were also no decisive correlative conclusions that could be drawn based on our data, therefore it would be unreliable to say that the data we gathered gives us a good grounds on which to conclude the unknown contents of the unknown dyes. Errors and suggestions: I am not sure exactly how the experiment was supposed to go or what the desired results were. There were some issues inserting dyes into the test gels. Also, there was some measurement confusions as well as a confusion as to exactly how much time the process would need to complete. In the future, I would say get more reliable samples, and have a better way or teach the testers how to inject the dyes to maintain accuracy and attain good or desired results in order to learn a lesson.