Lynna Ye - Week 2.2: Ware Lab

Last week, I started doing PCRs to amplify the cytochrome c oxidase I (COI) gene found in the mitochondrial DNA that I extracted from the dragonfly legs. The COI barcode has distinct differences in its sequence between species, so I will be using it to identify which species the larvae belong to by comparing their sequences to those of the adult specimens. Melissa, a grad student from Colombia, helped walk me through the procedure on Tuesday. After adding primers, Taq polymerase, etc. to my first batch of DNA, we set the temperatures and times on the thermal cycler, which subjects the DNA to specific periods of specific temperatures to allow the DNA to unwind, the primers to latch on, the polymerase to replicate the DNA, etc. in order to amplify the region that we want. Meanwhile, as the thermal cycler ran for approximately 2 hours, I continued doing extractions.

When the PCR was complete, we used gel electrophoresis,which uses an electric current to pull down the PCR product, to determine the size of the fragments I got from the reaction. Unfortunately, after running the gel we found that no distinctive bands could be seen. Melissa said that we probably have to adjust the temperatures, so the next day, I ran another PCR with the same DNA, with a few adjustments to the thermal cycler's program. However, the gel once more revealed unsatisfactory results.

The next day, we used different primers that would amplify a slightly longer portion of the COI gene and tried the PCR again, but alas, it was still unsuccessful. Also, the spectrophotometer that I was using began to give me odd and inconsistent results, so on Friday, I ran a gel instead on the extractions that I finished. Since I ran out of spin columns and the new extractions kits have not yet arrived, I cannot continue with extractions, so I spent the rest of Friday helping to label the dragonflies that Melissa and Will (another grad student) caught in their recent trip to Wisconsin. While it was a simple task, I found it fun to finally be able to touch some whole dragonflies, since until that point I have only been interacting with legs in tubes.

I am excited to see what this week has in store for me!