Second Week of ADHD Research

I had a wonderful day at the lab on Monday, June 18. First, I met with my principal investigator, Dr. Allison Ashley-Koch. We talked about an overview of my project. Unfortunately, the single nucleotide polymorphisms as well as the assays have not arrived yet. In addition, they were running low on ADHD DNA, so they have to wait until the DNA bank refills it before I could start on my project. As a result, I spent my day shadowing Chris, my mentor. This was a great introduction to me, attempting to learn more about the various instruments that they use in the lab. Moreover, with this practice, I will be more prepared when I start doing my own work on the ADHD DNA. Chris showed me how to use the Taqman recipe, which is used to figure out how much master mix, water, and assays should be added in accordance with the number of DNA samples. Next, I had to pipette the master mix onto the micro-well plates containing the DNA of patients enrolled in the Caregiver study. We then ran them on the PCR machine. After lunch, watched the Biomek robot transfer DNA into the micro-well plates. It was so fascinating to see how precise the machine worked. Lastly, we looked at the results from the Taqman plates earlier that day, and they were all quite successful.

Tuesday was another fun day in the lab. After speaking with Dr. Allison in the morning, I went to the third floor lab of the Center for Human Genetics Building. Chris asked me to pipette the Taqman recipe unto the micro-well plates containing DNA of patients enrolled in the Hostility study. However, before doing that I first had to make the recipe by adding water, master mix, and 20x assay. It was thrilling to do nearly the entire process by myself. After having to pipette two plates, I placed them in the centrifuge and took them upstairs to the PCR machine so that they can be ran for forty cycles. I was actually able to work the machine by myself (with instructions and Chris’ assistance of course =)). Then, I had to do the Taqman again, except this time, I used a different assay. This is because I was trying to find a different single nucleotide polymorphism (SNP) in the DNA. Upon returning from lunch, I created folders on the computer for my data to be stored in when we scan the plates. I also tried to find the gene and chromosome of the SNP that I was trying to observe. When I finished, it was time to take the plates out of the PCR machine so that they can be scanned using the 7900 machine. We then went to the computer to look at the data that we produced. None of the NTC controls were amplified, so we were able to submit the data to the clinical people for analysis. Finally, to end my day, we used the Biomek robot again. Overall, I was thrilled to be able to learn new techniques.

On Wednesday, June 20, I came to my lab and proceeded to the Post PCR refrigerator to get the plates that I prepared the day before. Chris allowed me to scan the plates by myself. Fortunately, I remembered most of the process that he showed me the prior. After scanning, he asked me to do the Taqman for the C-plates for Hostility. Since not all of the micro-wells contained DNA samples, I had to look at the chart to see where to pipette the Taqman recipe. I carefully completed that tedious process and centrifuged my resulting plates. Next, I ran the plates onto the PCR machine. When I returned after lunch, I did some more Taqman on DNA from patients enrolled in the Hostility study. Upon completion, I ended up with about fourteen plates. I went upstairs to store the plates in the cold room. This was a great day of practice and precision.

I had a rewarding day at the lab on Thursday. All of the plates that I ran on the PCR machine the prior were successful. I then looked at the results on the computer and none of the NTC controls amplified. After which, Chris showed me the DNA sequencing machine, wherein he uses a restriction enzyme to find a single nucleotide polymorphism in an insertion deletion. This is completed using micro satellites. He then asked me to do some more Taqman. I finished the list of assays needed to be run. Upon completion, I felt proud that I completed about twenty micro-well plates. Moreover, I am happy to say that my ID badge finally works, which means that I’ll be able to access most of the doors in the Center for Human genetics Building !!!

Finally Friday was more of an analyzing day. First, I scanned twelve plates unto the computer so that they can be run through on the 7900 machine. After a few minutes of waiting, I placed the plates in the cold room and analyzed the results to see if my PCR from the previous day was successful. To analyze the results, we used the program SDS 2.2.2. It generated graphs of the data, which is then submitted to a statistician for further evaluation. As it turned out, all of the plates that I completed were great. When I finished looking at the data that I produced, Chris showed me some useful tools in trying to find known mutations. There is a website called http://genome.ucsc.edu/ that is very helpful in doing so. We also used the Biomek robot to prepare for the PCR. Chris promised to show me how to use the PCR on Monday, so I’m very thrilled for that. Overall, this week was wonderful !!!!!