I'm trying to isolate spontaneous streptomycin resistant clones of Neisseria meningitidis. I have done this before, and I usually pick up mutations in rpsL (K43R or K88R) in the standard strain we use. With the strain that I currently work with I'm not able to pick up decent streptomycin resistant clones; what we find is very low level resistance and no mutations in rpsL. Does anybody know whether other mutations/ other antibiotic resistances may interfere? Or whether it's possible that bacteria contain more than one rpsL copy?
We finally managed to get StrepR mutants by transforming the strain with rpsL with mutation K43R from another strain. This didn't result in uptake of the DNA (the silent mutations present in the donor strain's rpsL were not present after transformation), but somehow did result in the isolation of some spontaneous mutants at 500 and 100 ug/ ml Strep. All of them contained the R43K mutation. No clue why the direct plating on Strep didn't work and this finally did...
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