How does the lack of antibiotic in LB affect the growth of antibiotic resistant - (Nov/02/2013 )
Hi, I have been searching for some information regarding my experiment but I cannot find any directly relevant. I am about to perform experiment to check whether the lack of ampicillin in growing medium will affect the growth of bacteria. Being more precise:
I am going to use E.coli. Two sets- the same strain transformed with two different plasmids respectively. Both contain plasmids with ampR, the difference is the size of the plasmids and the copy number. The bacteria have been previously grown on two respective agar plates containing ampiciillin (100 ug/ml). Now I am going to grow them in LB (no agar of course) and what I am going to do exactly is grow the ampicillin resistant strains in LB medium which does not contain ampicillin. I am also going to record the absorbance (bacteria will be in two separate erlenmeyer flasks at temp over 30 degrees in shaker) to monitor the bacterial growth. Absorbance will be checked every 20 minutes for approximtely 4 hours (at 550 nm). I have not performed this yet so have no data so far. I cannot find any similar research anywhere to compare my future results which worries me as I do not know what to expect and how to detect any errors. Hence my questions:
Does anyone know will the bacteria grow at all in this sort of non selective medium? If yes, will that be only these cells that did not take up plasmids or lost it? Will the absorbance readings will be relatively high or low, or will they be higher or lower than the abs readings comparing to when antibiotic resistant bacteria were grown in LB containing amp (I have performed this part already)? Does the salt concentration have anything to do with that? Does the plasmid type somehow affect the results? How do I know that something has gone wrong (too high, too low readings for certain strains, anything else)? Does anyone know what sort of predictable values of abs measurements are?
I know many questions and I am not expecting anyone to answer all of them, just trying to give you an idea of what I am trying to find out. I would be really thankful for any hints and also if anyone have any links to some similar research that would be great too.
E. coli without a plasmid (or with one) will grow in LB without antibiotic. Likely you will find that cells with plasmids grow faster without antibiotic, since the cells will not have to produce antibiotic resistance proteins before growth is permitted.
Thank you! That is very helpful.