Introduction: In this lab we used the polymerase chain reaction, or PCR for short, to amplify a piece of “Lambda Bacteriophage” DNA which is a virus that attacks bacterial cells. The PCR machine is a useful tool because it focuses on a region of DNA and amplifies a single molecule into billions of copies. Outside of this lab, the invention of the PCR has become an essential tool for testing DNA fingerprints, detecting genetic disorders and so much more in the world of genetics.
Results: Standard Values:Well Values:Gel On UV Light:(Unfortunately Julia and Sophie's results did not show up well on the gel because they made an error in their procedure) Graph:Conclusion: We successfully learned about the PCR and how to use it in this lab. PCR is a method of DNA replication that is able to take a small fragment of the DNA and make many copies with the help of a thermocycler. We tested this by using Lambda DNA which gave us notable results, especially when compared to the control lane. The purpose of the control is to show how the DNA travels without any manipulation from the thermocycler. The control sample traveled 1.23 cm which was significantly less than the rest of the samples (and resulted in 21,500 basepairs). This proves that without the thermocycler, there would not be accurate results, and that thermal properties are important in the use of the PCR. Although we had a successful experience with the PCR, I wonder if there are other methods that can accomplish this same process? If so, what is more efficient? Also, how are primers designed to target specific locations of the DNA strand? This lab was very interesting to me especially since we were able to work with real DNA! I also understood how the procedures and techniques we learned in the first lab, are extremely important to practice when working with DNA. I look forward to learning about more lab procedures, as well as continuing to work with DNA! Discussion: PCR artificially simulates DNA replication in a cell, through many cycles and temperatures. It uses primers to focus on a certain region of the DNA strand to amplify it. There are three steps involved in the PCR, denaturing, annealing, and extension. The denaturing step is supposed to represent how the helicase enzyme in the cell breaks apart the hydrogen bonds in the DNA strand. In the PCR, instead of using enzymes to break the bonds, this is accomplished by using heat. The next step is annealing. In DNA replication in the cell, RNA primase primes each nucleotide on the DNA template. This is similar to the PCR, where there are synthetic primers that “anneal” to the DNA template. However in PCR, there are two primers used for each DNA strand. The final step is extension. The primers laid out by annealing serve as the starting point for extension. Taq polymerase binds to each of the PCR primers to add nucleotides. Taq polymerase acts like human DNA polymerase because it adds nucleotides in one direction. This is why it’s necessary to have two primers on both strands of the DNA. Taq polymerase comes from the bacterium “Thermus Aquaticus” which originates from hot springs. Because it lives in hot springs, it thrives at high temperatures, which makes it the most useful for PCR since the denaturing step is primarily done through thermal properties. DNA is naturally colorless, so if we were to run a gel with just the DNA, we wouldn’t be able to see the distance it had traveled. This is why we use a loading dye when putting the samples in the wells to ensure that we can see our results under the UV light after electrophoresis. The known size of the amplicon in this lab is 1,106 base pairs. Our closest value to this was Sarah’s value which was 1,475 base pairs. The reason behind why this is slightly inaccurate is because I graphed our values by hand and had to estimate the base pair value. Bonus: Works Cited:
Study.com, Study.com, study.com/academy/lesson/pcr-synthesizing-dna-using-polymerase-chain-reaction.html. “The Role of Taq Polymerase in PCR.” Sciencing, sciencing.com/role-taq-polymerase-pcr-7298417.html.
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ABOUTHere are updates from my Genomics class.
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