A mutation in Escherichia coli K12 giving resistance to about 5 μg/ml of streptomycin was found to be cotransducible by P1 with proA and proB, and is located at about 8·5 min on the chromosome map. The locus is named strB. A second mutation to the same resistance level was not cotransducible with either proA or proB and must be located elsewhere. Both mutations cause a marked increase in R-factor mediated streptomycin resistance, and significant decreases in resistance to several other antibiotics, both in the presence and absence of an R-factor determinant for the same antibiotic. The two mutations differ in their effects on bacterial sensitivity to crystal violet and EDTA.