SUMMARY An Hfr strain G I I and an F- strain D I of Escherichia coli K 12 both containing the R-factor R1a were treated with ethyl methane sulphonate (EMS). Mutation of the bacteria to high ampicillin resistance occurred at a frequency of IO-~ to I o-~ and it was coupled with a simultaneous increase in resistance to chloramphenicol, streptomycin and sulphanilamide. Resistance to all the antibiotics was infective. Increased ampicillin resistance was due to increased penicillinase activity of the bacteria. In a strain containing a mutant R-factor (RI BI) this was due to an increased quantity of an enzyme that seemed to be identical to that of a strain con- taining the unmutated R Ia. Metabolism of chloramphenicol in strains containing R I B I was greater than in those with R Ia. Mating experiments with an Hfr strain carrying R Ia or R I B I revealed that the two R-factors reduced fertility to about the same extent. Pair formation was two to three times greater in the presence of R I B I than in the presence of R Ia, and R-factor transfer was increased more than chromosome transfer. It is likely that the RI~ genome is read two to three times more efficiently in strains containing the mutated KI BI, perhaps due to an increased number of copies of the R-factor genome per bacterium.