Since few months, I have started working in bacterial bioflocculation experiment with 5 screened strains. I've successfully characterized the first strain; whereas 4 other screened strains are stored under ultra-low temperature. This unpredictable behavior started from the next strains under characterization process. The polymer/product concentration as well its activity started varying in each time of experiment get repeated. So, I’m wondering how this happened with my screened bacterial strains and when things were done so precisely in every step? If anyone can explain this and suggest if something has been lacking or been overlooked in the process, I would greatly appreciate it.
The physiological state of your cells would greatly affect your observed results. Try to ensure that you use cells at the same phase of growth in all experiments. If you use cells that are in the stationary phase now and use those in the log phase later, your product concentration would definitely vary. I suggest you first conduct growth curve experiment for each strain so as to know the time each reaches log phase, etc. You should then grow each strain till the late log phase for example and always use that in all the experiments. This is not ruling out other possible sources of variation, but in my experience, this affects the consistency of results a lot.
May we ask what strains you are working with and what polymer/compound you are extracting?
Did you immediately make frozen stocks of your strains when you first received them or did you just store them in low temperature conditions?
From my experience, bacteria are unique, sometimes they grow, sometimes they don't. Sometime that make the compounds we want to study and sometimes they don't. My suggestion is to always keep backups. I once had a bacterium that stop making its compound and I spent months going back to my frozen stocks to get it to make the compound. Well, it never did, I guess it got acclimated to the lab and stopped making it. As a last resort, I plated out some old R2B test tubes from my advisor's work station containing the organism when she first isolated it (these were sitting at room temperature for at least 2 years), and guess what? I got the organism that made the compound back. Weird? Bacteria are interesting organisms.
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