Flac mutants have been isolated in Escherichia coli K-12 which carry dominant mutations resulting in insensitivity to transfer inhibition by the Fin(+) I-like plasmid R62. These mutants were still sensitive to transfer inhibition by the fin(+) F-like plasmid R100 and, conversely, FlactraO(-) and traP(-) mutants, which are insensitive to R100 inhibition, were still sensitive to R62. The sites of action of the two inhibition systems are therefore different. Furthermore, inhibition by R62, unlike R100, did not require an F-specified product. Like R100, R62 prevented transfer, pilus formation, and surface exclusion and, therefore, probably inhibits expression of the transfer operon traA through traI. However, R62 was different from R100 in inhibiting transfer of J-independent mutants, indicating that its effect on the transfer operon is probably direct rather than via traJ. This is consistent with the different sites of action of the two inhibition systems. None of the Flac mutants overproduced pili in the absence of R62, although one mutant differing from those described above showed increased levels of transfer and surface exclusion.