How would I do this experiment? [Advanced]

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How would I do this experiment? [Advanced]

Post by fstevens » Sun Nov 16, 2014 11:10 pm

This a question about how to do a experiment. I am a beginner so how to do each procedure would be very helpful!
Things to know: my microscope is up to 2000x. I am a beginner biologist so please do not speak in very technical terms.

My experiment: Testing Kefir (a friendly 'probiotic' bacteria drink) vs. Escherichia coli (bought from Carolina Scientific- Living, K-12 Strain, Tube) I want to test how well Kefir can kill off Escherichia coli (or vice versa). What I want is to introduce around the same amount of E coli cells to the same amount of probiotic cells. I am going to test different store-bought brands and home made Kefir. Is there some way to count the bacteria and measure the exact amount? Can I do this only with a 2000x microscope? It would be very helpful to get full instructions how to do this, and I would learn a lot. Here is one more simple overview of my experiment:

-I want to find out which type of kefir is best against Escherichia coli.
-I am buying the e-coli from ... / if this is a bad place to buy it, let me know.

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Re: How would I do this experiment? [Advanced]

Post by Hrushikesh » Mon Dec 15, 2014 5:00 pm

Have you planned how shall you count E. coli cells and Kefir cells and distinguish between them?
If you use turbidity as a measure of concentration of bacteria, it may not distinguish between those two, moreover you may get noise due to killed/dead cells in the readings.
If you use live cell stains, and then use microscopy to count cells, then you may be able to do it, but how shall you selectively stain E.coli or Kefir cells to be able to count them separately? If you find a selective stain for E.coli you may be able to count them separately.
Also, what you can do is, use serial dilution technique and then pick up colonies on final plates and distinguish as Kefir or E. coli based on microscopic observations and possibly some tests.
Does this solve your problem?


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Post by JackBean » Fri Dec 19, 2014 2:46 pm

As you must be able to grow the bacteria (otherwise you cannot perform the test at all), it would be better to perform dilution of your samples and spread aliquots of such on plates with growth agar (for E.coli simple LB, for Kefir, no idea). Let them growth (E.coli for 1 day at 37°C) and count number of colonies (pick plates with 10-300 colonies, that's why you do the dilution, because <10 colonies makes big error and hundreds of colonies are hard to calculate (for that, split the Petri dish into several compartments with pen and count each separated)). Also, make each dilution in replicates, two or three at least.
If the respective bacteria will have different morphology of colonies or even will require strictly different nutrients/conditions, you can use this method also for distinguishing between the two.

Cis or trans? That's what matters.

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