Alu PCR Lab
Purpose
The purpose of this lab was to be able to successfully isolate DNA and prepare a PCR reaction for amplification of an Alu insert. With our results, we were to determine whether or not we carried the frequency of the Alu insert in our DNA.
Hypothesis
If we use PCR to determine how frequent the Alu insert is in my DNA, then I predict that I will have a heterozygous allele.
The purpose of this lab was to be able to successfully isolate DNA and prepare a PCR reaction for amplification of an Alu insert. With our results, we were to determine whether or not we carried the frequency of the Alu insert in our DNA.
Hypothesis
If we use PCR to determine how frequent the Alu insert is in my DNA, then I predict that I will have a heterozygous allele.
Procedure
Results/Conclusion
According to the gel samples, my genotype was heterozygous, meaning one band was at 415 bp and a second band was at 715 bp. This means that my hypothesis was correct. From this lab, I learned the importance of reading instructions completely and clearly. The experiments could have easily gone wrong since there were so many precise, small measurements that had to be taken to receive accurate readings.
- Swirl 10 mL of saline solution (0.9% NaCl) in your mouth for 30 seconds and expel into a cup.
- Transfer 1000 micro liters of the suspension into a labeled 1.5 mL microfuge tube.
- Spin the tube in a microcentrifuge for one minute to pellet the cells.
- Pour the separated supernatant into a cup, leaving 1 mL on top of the cell pellet. Mix the remaining solution by racking it.
- Withdraw 50 micro liters of the solution and add it into a tube of 5% Chelex.
- Place the Chelex microfuge tube into a heat block for 10 minutes. Once the solution is heated, place in the centrifuge for 1 minute.
- Withdraw 50 micro liters of the solution and place it into a tiny PCR tube.
- Add 20 micro liters of Primer Mix and Master Mix to a separate tube. Then add 10 micro liters of the DNA solution to this tube.
- Add 5 micro liters of loading dye to the PCR tube.
- Load the contents of the PCR tube into the agarose gel tray.
- Attach electrodes to the the power supply and the agarose gel and check the samples for 25-40 minutes.
- Check for results.
Results/Conclusion
According to the gel samples, my genotype was heterozygous, meaning one band was at 415 bp and a second band was at 715 bp. This means that my hypothesis was correct. From this lab, I learned the importance of reading instructions completely and clearly. The experiments could have easily gone wrong since there were so many precise, small measurements that had to be taken to receive accurate readings.