The transposable element Tn5 (Kan) is being used as a mutagen in E. coli K-12. Although the more than 300 Tn5-induced auxotrophs isolated are due to insertion into a large number of genes, some preferred regions have been detected. In most insertion mutants Tn5 is able to excise perfectly, restoring the cell to prototrophy. Extended deletion mutants are isolated and studied using a strain in which a thermoinducible lambda, deleted for lambda att, and carrying Tn5 has been integrated into an ilvA: Tn5 strain of E. coli. Thermoinduction of a rec minus derivative of this strain yields many delection strains. The Tn5 insertion mutants are also proving useful for the determination of operon polarity and for the detection of internal promoters.