Blogs

These past two weeks have been so busy, as I'm not only trying to finish up my lab work but also work on a presentation and paper for my internship.  As usual, the majority of my lab work has been running PCR tests and cloning samples when necessary.  

This week was not as busy as previous weeks. I did not have to run time-points like the previous weeks, and did not grow Pseudomonas putida. Most of what I did was to wrap-up.  I finished the measurements from last week’s experiment, finished analyzing last week’s data, ran an experiment for catalase activity with different concentration of catalase, edited my PowerPoint for the final presentation, and finished up my final paper for this internship.

On Monday morning I took photos of my Carbon Source “Feeding” Experiment. I noticed that the biofilm in lignin appeared to be somewhat weaker than it was before and the biofilm in humic acids was much thicker. This observation lead me to believe that P.putida cannot consume any more lignin, but is still digesting the humic acids. My mentor and I noticed that the solution in the oxidizing culture of humic acids (with the thick biofilm) was a much clearer solution than the non-oxidizing mutant in humic acids.

There’s only one more week left for this internship. Everyone in the lab was so busy. It could be that everyone was wrapping up their experiments. This week I repeated the experiment from last week but with different concentration levels of Iron: 1.8 and 9 uM Fe. The measurements were the same: catalase activity, hydrogen peroxide concentration, the presence of pyoverdine, dissolved and particulate manganese (Mn), manganese oxide (MnO_x), and the culture density.

This week I finished up my final round of batch experiments with a chemical called 1,3-dinitrobenzene. It was pretty quick and easy, I finished it, plus a couple extra TNT experiments within the week. DNB is faster than everything except for TNT. 

This is a graph of the final product of my experiments: 

Monday: More preparations for the final paper, had a professional development meeting regarding posters for presentation, and had a meeting with Charles and Tuomas.

Tuesday: Wrote more for the final paper and had final meeting with Charles about code.

Wednesday: Wrote more for the final paper and had final meeting with Tuomas about code.

This week I started cloning. The first thing I did was preform a normal touchdown PCR of the sediment samples using the bacteria primers. Next I made LB agar plates for culturing later on in the process. I then preformed a purification step on the PCR product in order to get the extra protein and enzymes out before preforming the TOPO cloning process and freezing the product for this week when we will continue the process.

On Monday I did not get as much information from my motility plates as I would’ve liked, but I did get a few good photos. There was denser and darker oxidation occurring next to certain carbon sources. More oxidation means that there are more cells, and if there are more cells, there must have been more growth in that area. Another way to observe the bacteria seeking and eating the carbon source was when there was a bulge in the circle towards a certain carbon source. This means that the bacteria made flagella so they could swim more towards the food source they could eat.

Everything is becoming so busy now. The summer started off at an easy pace and now the stress is beginning to set in. Most days I find myself so busy with things that I don't realize how late it is. The new barcoded primers came in earlier than expected. So, this week Rick and I have been trying to determine the best method of barcoding the samples. So far, things haven't been working too well. I did a PCR with the barcoded primers today and they didn't amplify the DNA.