Purpose
The purpose of this lab was to become more familiar in a lab setting and to learn about PCR though our alu repeats.
Hypothesis
Before I went into the lab my hypothesis was that my DNA alu repeats would show that I am from European descendant.
Procedure
The procedure of that we used for this lab was from the Bay Area Biotechnology Education Consortium (BABEC) from their Alu PV92 PCR report.
Data/ Observations
The first day we were in the lab I noticed that there was a little cell pellet on the bottom of the tube. The cell pellet was mostly a clear white and you could almost see little circles inside the cell pellet. I noticed that the Chelex was clear little spheres that rested at the bottom They were able to move freely when the tube was tilted.
The second day that we worked on the PCR lab I noticed that we were working with very small amounts of liquid. I observed that whenever we wanted to have the liquid resting at the bottom of the PCR tube we had to centrifuge the DNA filled sample because it brought the drop of liquid to the bottom of the tube.
On the third and final day of being in the lab I noticed that you had to be very precise when setting DNA and dye into the gel. My other observation was that the gel was very pigmented and that when it was placed in the gel mold it spread over the gel.
The second day that we worked on the PCR lab I noticed that we were working with very small amounts of liquid. I observed that whenever we wanted to have the liquid resting at the bottom of the PCR tube we had to centrifuge the DNA filled sample because it brought the drop of liquid to the bottom of the tube.
On the third and final day of being in the lab I noticed that you had to be very precise when setting DNA and dye into the gel. My other observation was that the gel was very pigmented and that when it was placed in the gel mold it spread over the gel.
(Figure 1):
2% agarose gel ran at 150v for 20 minutes and stained using gel red for 72 hours land 1a and 2a have 100 bp ladder. Lane 1 b-h and 2 b-h have 20 micro liters of 50 micro liters of DNA and 10 micro liters of loading dye solution. Lane 1g had the sample I worked with.
Analysis
We were unable to look at our alu repeats to see our ancestry background because not enough people had data so we could not proceed further into the lab to observe our ancestry, but had we had the chance my hypothesis still stands; I think my alu repeats show European descent. Since we had no class data we used numbers to simulate results. We did this so we could practice doing the math to calculate Geno type frequencies. Errors and mistakes most likely occurred when we were rushing to to finish before the end of the period. This lead to errors because we threw accuracy away when rushing. Mistakes and errors probably also happened when the directions were unclear to us. Things we could change next time we do this lab is actually having crushed ice for steps 1-4 under the Polymerase Chain Reaction section because it would have been easier than working with ice cubes and it would have been easier to keep the tube cold when ice was fully surrounding it. Another thing that could have gone better was having more time in the lab because everything was rushed and that led to more room for error. This lab leads me to want to investigate my ancestry because we were unable to investigate it in the lab.
(Figure 2):
This was the math we had to do to find the Geno type Frequencies
Conclusion
PCR is challenging when pressed for time. In this lab, we learned about PCR through gels by using our DNA. What made the lab so difficult was that we had to be very precise when measuring the saline, cell sample, master mix, chelex, and the other liquids. Other parts of the lab required you to only take liquid from the surface or the bottom of the test tube. We also had to very conscience of the temperature of the PCR tube because some parts had to be strictly on ice and other parts had to be at an exact temperature for exactly 10 minutes. PCR is difficult given that statistically 83% of the class did not have any results. This lab was so hard because everything had to be precise and it was our first time in the lab so everything was brand new which made every concept foreign. My conclusion based on my data, evidence, and personal experience is that PCR is hard.