Weird thing: Cml resistant S. aureus strain (RN4220) - (Jun/05/2007 )

Our lab got a S. aureus strain RN4220 ( - mec gene) from another lab as a gift and this strain does not have vector inside. To test if the strain is contaminated, I grow RN4220 in TSB medium containing Ampicillin (100ug/ml) or Chloramphenicol (Cml) (5ug/ml). Results show that RN4220 cells cannot grow in the medium with Amp or Cml. However, RN4220 grown in TSB overnight also grows very well on the TSB/agar plate with Chloramphenicol (5ug/ml).

Does it mean that our RN4220 strain gains resistance to Chloramphenicol? I checked NARSA web site and didn't find any RN4220 strain resistant to Cml. What's wrong? I want to use Cml to select positive colonies during transformation.


Thank you for your suggestion.

I always use 10ug/mL chloramphenicol to inhibit wildtype RN4220 growth. Additionally, since chloramphenicol is solubilized in ethanol, it often is not equally distributed in agar (floats), is more likely to precipitate in TSA, and generally results in lower concentrations when made into plates (forms "pockets" rather than even distribution). Watch as you add it to your agar -- you have to be careful to watch for precipitation. It's normally fine in liquid, particularly if you incubate with shaking.

My guess is that your plates simply have too low of a chloramphenicol concentration to inhibit the growth. I'd raise it up and see if that takes care of it.


Amanda