An msDNA operon, consisting of genes for msDNA and a reverse transcriptase, is present in Escherichia coli B and absent from E. coli K12. We have found that the msDNA operon is located on a DNA fragment, longer than 15kb, that is absent from E. coli K12. Using conjugation, P1 transduction, and nucleic acid hybridization between E. coli B and E. coli K12 strains, we have located the position of the msDNA operon on the E. coli B chromosome at a site that corresponds to minute 19 on the genetic map and to position 900 on the physical map of the E. coli K12 chromosome.