Seven citrate-utilizing (Cit+) Escherichia coli strains originated from horeses (2strains) and cattle (5 strains) were examined for transferability of Cit+ character and the presence of bacterial plasmids. All 7 strains harboured plasmids in the cell, although the Cit+ determinants in the Cit+ E. coli strains tested could not be transferred to E. coli K-12 strain. Two horse strains possessed about 60 megadaltons plasmids; the plasmid in a strain OH3026 was thought to be non-conjugative Cit plasmid, but that in OH3025 was not exactly shown to encode the Cit+ functional ability. Plasmid DNAs prepared from 2 strains (OH3035 and OH3036) of 5 bovine strains could produce Cit+ transformants, but no Cit+ transformant was obtained in the remaining 3 strains. The representative non-conjugative Cit plasmid, pOH3035 (20.5kb), prepared from the Cit+ transformant of pOH3035 was examined by restriction endonuclease, and its EcoRI-cleaved B fragment (5.5kb) carrying Cit+ functional gene was cloned to a vector plasmid pBR322. The Cit+ recombinant plasmid, pOH8 constructed by cloning was analysed for potential transposability of Cit+ gene, but the transposable element coding for Cit+ determinant was not obtained from pOH3035 and pOH8 in this study. The restriction map of pOH3035 was also compared with that of a plasmid pBR322 carrying citrate utilization transposon, pOH2, and the relative similar order of recognition sites for BamHI and BglII enzymes was found on Cit+ functional gene of the 2 plasmids.