S-STEM Scholar Alex Hugenberg-Cox Blog

      This week we discussed the math behind our qPCR analysis, and learned the steps to do it ourselves. While we do get the calculations from a computer, its always good to be able to double check the calculations, and to be able to find where an error is if their is one. We covered the calculations for relative expression (called the livah mathod), not absolute expression as that's the calculations used in our project and also used for most qPCR calculations.       After covering how to do the calculations, we also discussed and reviewed our procedure. We hadn't been getting no significant change in gene expression when under oxidative stress from H2O2. Even when tracking the KatA gene, which should have light up with H2O2, we saw no significant change. When looking at the paper we were basing our procedures on, we saw that they actually halted the H2O2 reaction by adding catalase. After a small discussion, we decided that next week when moving forward with our procedure we

  Last week we decide against continuing with pfs and instead using both LuxS and KatA. While we didn't get to qPCR this week, I did get to perform RNA isolation by myself and and do a new procedure: alcohol precipitate.  We started the week on Monday with a graham stain. All our cells looked good, and we had a ton of D.rad to work with.  Running the RNA isolation on my own was difficult. There was alot of back and forth, and i was constantly leaving and re-entering the hood. Deja was able to arrive in time to help me for the last few steps, but for the first hour and a half I was doing what we usually have two people do. Since we do the RNA isolation in the biological hood, I was very worried about contamination or even just forgetting a step trying to do everything.  I did have a hiccup during one of the steps. I had been given instructions to try to do everything possible in the -20 ice box, instead of keeping it in normal ice when possible but doing normal steps in the hood in