I want to control bacteria growth at a gradient density to confirm whether the antibiotic production is density-dependent. But I don't want possible effects of culture medium during growth. For example, what is the threshold population density that the bacteria will start to produce antibitics? I know continuous bioreactor might be an option but not sure whether there is limitation for constructing a density gradient.
I am not sure I understand your question, but I'll give it a try. Grow your bacteria to an optical density of 0.6 - 0.9. Store this at - 80 in glycerol (20% v/v). Determine what CFU/ml you have stored by plating out serial dilutions. Once you know what cfu/ml you have, run your experiment with a different starting inoculmns (CFU/ml) that you will prepare by diluting your stored master culture. Using CFU/ml form low to high concentrations i.e., 1 x10^4, 1 x 10^5 etc under your conditions should allow for you determine if the response is density dependent.
I am not sure I understand your question, but I'll give it a try. Grow your bacteria to an optical density of 0.6 - 0.9. Store this at - 80 in glycerol (20% v/v). Determine what CFU/ml you have stored by plating out serial dilutions. Once you know what cfu/ml you have, run your experiment with a different starting inoculmns (CFU/ml) that you will prepare by diluting your stored master culture. Using CFU/ml form low to high concentrations i.e., 1 x10^4, 1 x 10^5 etc under your conditions should allow for you determine if the response is density dependent.
Hi, Daniel.
First of all are you sure that your bacteria produces "antibiotics"? More than density dependent the growth phase might also be an important factor to take into consideration. If you take the Penicillium example, this fungus only produces antibiotics in the stationary phase.
Another that might be important is the richness of your culture media. Sometimes, the poorness of the media helps to turn on some genes that are not expressed when the conditions are the best for their development.
Now, to what concerns to your question, I totally agree with Shane. First you have to determine your CFU/ml only then can you start to test your bacteria at different concentrations. If you want to avoid any inhibition from the accumulation of metabolites I think that if you try to refresh your culture by adding new culture media might help to solve this question.
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