Summary: R68.45 and other similar broad-host-range (IncP) plasmids carrying a tandem repeat of the 2.1 kb insertion element IS mobilize the chromosome of many different Gram-negative bacteria. To analyse the structure of R68.45-chromosome cointegrates, whose involvement in the mobilization process had been postulated previously, we selected for the stable integration of R68.45-like plasmids into the chromosome. Two plasmids were chosen: pME28, a transfer-deficient, mobilizable RP1 derivative with an inactive replication control () gene, and pME487, an R68.45 derivative with a (ts) mutation causing temperature-sensitive replication. Chromosomally integrated pME28 and pME487 were found to be flanked by single IS elements. This structure is in agreement with a “cut-and-paste” mode of R68.45 transposition. pME28 and pME487 showed a low specificity of insertion but rarely (<0.1%) induced auxotrophic mutations. Hfr (high-frequency-of-recombination) donors of could be obtained by chromosomal integration of pME487 or pME28; in the latter case, the transfer functions lacking from pME28 had to be provided on an autonomous plasmid. Hfr donors gave higher conjugational linkage and transferred longer stretches of the chromosome than did R68.45 donors. This suggests that the integration of R68.45 into the donor chromosome is short-lived in


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