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Hydrogen Peroxide and Pseudomonas Purida - Week 4

This week was all about hydrogen peroxide(H2O2) . The H2O2/amplex red protocol, which I have been practicing, was used to measure the H2O2 concentration in a minimal medium with bacteria called pseudomonas putida GB1.  We want to test if the bacteria would produce H2O2. So far we only have collected six time points. My mentor, Matt and I planned to take another point on this coming Monday.

At the beginning of this week, minimal medium A and LB medium were prepared to grow bacteria (pseudomonas putida GB1). On Monday, one colony of P. putida was inoculated from one of Christine Romano’s plate into a liquid LB medium. This media has enough nutrient for them to grow healthy and happy overnight. Then on Tuesday, we inoculated the overnight LB medium into minimal medium in eight different 125 mL flasks. Two of them were control with no P. putida in it: one with aluminum foil wrapped around it, and one without foil. The other six flasks were replicates with P. putida: three of them were wrapped with aluminum foil and three without. At the same time when I added the P. putida, I run a H2O2 assay. This was my first time point, T0. After four hour, I run the assay again and that was my T1. On Wednesday, I took two other time points, T2 and T3. I graphed all the time points fluorescence vs. time. The fluorescence I measured from the assay was decreasing at each time point, and it was lower than the calibration curve I did with the tested sample. Matt suggested that it could be because the P. putida was changing the minimal medium chemically. I still haven’t fully analyzed my data yet. I would have all my time points until this coming Monday.