Alu PCR Lab (Polymerase Chain Reaction)
Purpose: To learn to use PCR by trying to find an alu insert in our DNA
Purpose: To learn to use PCR by trying to find an alu insert in our DNA
Materials:
Cheek cells Chelex Master mix Primer mix TBE Buffer Agarose Gel DNA Stain Solution |
Centrifuge
Electric Scale Gel casting tray Tape |
Procedure:
1. Swirl 10 ml of saline solution in your mouth for 30 seconds. Spit out saline into a cup and swirl to mix cells.
2. Transfer 1000 microliters of the saline/cell suspension into your labeled microfuge tube. Spin in a microcentrifuge to pellet the cells. Pour off the supernatent, allowing 100 microliters to cover the cell pallet. Rack the sample.
3. Withdraw 50 microliters of your cell suspension and add it to a tube containing Chelex.
4. Apply Chelex tube to a heat block for 10 minutes.
5. Remove Chelex tube from heat block. Use a P-200 to withdraw 50 microliters of supernatent from the Chelex tube and transfer to a fresh tube.
6. Obtain a tiny PCR tube and keep on ice.
7. Pipette 20 microliters of Master Mix into the PCR tube. Then add 20 microliters of Primer Mix.
8. Add 10 microliters of your extracted DNa into the PCR tube.
9. Place reaction into a thermal cycler.
10. Retrieve PCR tube and spin in a microcentrifuge. Then, add 5 microliters of loading dye.
11. Create and pour gels. Add 1XTAE solution.
12. Load 15 to 20 microliters of the DNA/loading dye mixture into a well in your gel.
13. Load 5 to 10 microliters of a 100 base-pair ladder (molecular weight marker) into the one well in each gel for later comparison.
14. Run gels.
My groups results. Only some of us had the Alu gene
Results: My results shows that i'm +/-, which means i'm a heterozygous.
Conclusion:
While learning about my DNA was interesting, I believe that the most important part of the lab was learning about proper biotech procedures. The class made a mistake in the first part of the lab, and unfortunately had to start over. Although this was a bummer, we learned that this is frequent in biotech experiments, and it is important not to give up on the lab. We also got to practice using laboratory materials such as centrifuges, microfuge tubes, and micro pipets. Overall, this was a great way to learn about safe lab practices, proper use of equipment (balancing the centrifuge), and PCR.
1. Swirl 10 ml of saline solution in your mouth for 30 seconds. Spit out saline into a cup and swirl to mix cells.
2. Transfer 1000 microliters of the saline/cell suspension into your labeled microfuge tube. Spin in a microcentrifuge to pellet the cells. Pour off the supernatent, allowing 100 microliters to cover the cell pallet. Rack the sample.
3. Withdraw 50 microliters of your cell suspension and add it to a tube containing Chelex.
4. Apply Chelex tube to a heat block for 10 minutes.
5. Remove Chelex tube from heat block. Use a P-200 to withdraw 50 microliters of supernatent from the Chelex tube and transfer to a fresh tube.
6. Obtain a tiny PCR tube and keep on ice.
7. Pipette 20 microliters of Master Mix into the PCR tube. Then add 20 microliters of Primer Mix.
8. Add 10 microliters of your extracted DNa into the PCR tube.
9. Place reaction into a thermal cycler.
10. Retrieve PCR tube and spin in a microcentrifuge. Then, add 5 microliters of loading dye.
11. Create and pour gels. Add 1XTAE solution.
12. Load 15 to 20 microliters of the DNA/loading dye mixture into a well in your gel.
13. Load 5 to 10 microliters of a 100 base-pair ladder (molecular weight marker) into the one well in each gel for later comparison.
14. Run gels.
My groups results. Only some of us had the Alu gene
Results: My results shows that i'm +/-, which means i'm a heterozygous.
Conclusion:
While learning about my DNA was interesting, I believe that the most important part of the lab was learning about proper biotech procedures. The class made a mistake in the first part of the lab, and unfortunately had to start over. Although this was a bummer, we learned that this is frequent in biotech experiments, and it is important not to give up on the lab. We also got to practice using laboratory materials such as centrifuges, microfuge tubes, and micro pipets. Overall, this was a great way to learn about safe lab practices, proper use of equipment (balancing the centrifuge), and PCR.