Alu PCR lAB
My name is Aidan Evans, and I performed my research at San Marin High School. We started this project on Wednesday, August 26, 2015. We ended this project on Friday, August 28, 2015.
The purpose of this lab was to use PCR (Polymerase Chain Reaction) in order to view our DNA. We were trying to figure out if we had heterozygous, or homozygous genes. If we saw multiple bars of DNA on the gel, it meant we had heterozygous genes. If there was only one bar, it meant we had homozygous genes. To be honest, I cannot make a hypothesis based on what I already know. I am not yet familiar enough with DNA and my genes. If I had to guess, I would say I am heterozygous, because it sounds like it would be more common. That by the way has no meaning what so ever, so if I get it right, lucky me and if I don't, now I know. Our materials were: Cheek cells, chelex, the master mix, which consisted of nucleotides, and DNA polymerase. We also had the primer mix, which consisted of TBE buffer, and agarose gel. Our last material was a DNA stained solution. Procedure: 1. Vigorously swirl 10mL of saline solution in your mouth for 30 seconds. 2. Expel saline into a cup and swirl to mix the cells. 3. Label a 1.5mL microfuge tube with your PIN. 4. Transfer 1mL to 1.5mL of the saline / cell suspension into the labeled microfuge. 5. In a microcentrifuge, spin your saline cell suspension for 1 minute to pellet the cells. Be sure to use another student's sample as a balance. 6. Pour off the supernatant into your cup, being careful NOT to lose your cell pellet. 7. Rack or flick tube to mix,which will "resuspend" the cell and make an evenly mixed solution. 8. Obtain a tube of Chelex from your instructor. Label with your PIN. 9. Withdraw 50 micro liters of your cell suspension from step 7 and add it to the tube containing Chelex. 10. Put into 99°F heating block for ten minutes. 11. Take the tube out of the heating block, then open it to release the pressure. Close it back up, rack it, and then put it back into the spinner for 1 minute. 12. Obtain another clean microfuge tube and label it with your PIN. Also, write "DNA" on this tube. 13. Remove 50 micro liters of supernatant into the new tube. Do not get any of the Chelex pellets in it. PCR: 1. Label a PCR tube with your PIN number. 2. Pipet 20 micro liters of Master Mix into your PCR tube. 3.Change your pipet tip and add 20 micro liters of Primer Mix into your PCR tube. 4. With a new pipet tip, add 10 micro liters of your extracted DNA into your PCR tube. 5. Place your reaction into the thermal cycler and record the location of your tube on the grid provided by your teacher. After completing all of the steps and looking at the gel, I noticed that my section, although hard to see, had three bars. my section was second from the left, and as you can see, there are three "bands". What does this mean? As I said before, one who has heterozygous genes would see multiple bands, and one who has homozygous genes would only see one band. Using this piece of information, it is safe to conclude that I have heterozygous genes, because three bars is more than one. We were trying to figure out how to see our DNA using PCR (Polymerase Chain Reaction). We wanted to see if we had heterozygous genes, or homozygous genes. My hypothesis was that my genes would be heterozygous. My results agreed with my hypothesis. I saw three "bars" in my section, therefor I was heterozygous, not homozygous. |