Week 2: No Lifeguard on Duty

I was more or less on my own this week as Pete was going to be gone for a wedding. And though it was a little nerve racking to be on my own, there was a lot to be done.

Most of the week was spent doing DNA extractions, PCR and running gels. We will be looking at the DNA more closely next week so that we can verify that the protists we are interested are present in the water samples. I think I am starting to get better at DNA extractions; the first time alone was tough because I needed to keep referring to the protocol to make sure I was doing everything right. The second time around still took a long time, but I had a better grasp on the procedure so it went by a little faster.

I have been having some problems with PCR on the samples as only a few of them amplified. Sometimes there are inhibitors of PCR in the DNA extractions and therefore diluting the samples would make it easier for PCR to work, so I tried diluting the samples hoping that they would now amplify in PCR. This only worked for a few of the samples so next week I will be doing a re-extraction in order to purify the DNA even more. For the samples that did amplify I did a PCR cleanup on the samples so we can work with this DNA later.

This was a pretty busy week and there are still so many questions that need to be answered. I am really curious as to why these samples have such high DNA concentrations but are not amplifying well in PCR hopefully the re-extraction will help us produce better results. Also Pete will be back next week so I am excited to perhaps start some cloning experiments and do qPCR.