Segregation of Col Ib and drd7 into minicells.

The wild-type plasmid ColIb and its mutant drd7 derepressed in conjugation were transferred to Escherichia coli K12 P678-54 which produces minicells. Fertility functions of drd7 remained derepressed in the new host. P678-54drd7 transmitted the plasmid at a high frequency (28.6%) and it was effectively lysed by the phage If1. Significant amounts of 3H-DNA segregated from P67854Col+ into minicells dependent upon the presence of the plasmid. The depressed plasmid segregated more effectively into minicells than the wild-type plasmid. ColIb segregated into 2% whereas ColIbdrd7 into 8.4% of minicells. The difference in the frequency of segregation of the wild-type and the derepressed plasmid indicated different cell membrane attachment sites of each plasmid studied. Mini-drd7 were able to transfer the plasmid to E. coli Row at a low frequency (0.1%). Minicells carrying either of plasmid were capable to synthesize RNA and protein. RNA and protein synthesis were plasmid specific and the precursors were not incorporated into minicells without plasmids. Rifampin and chloramphenicol inhibited RNA and protein synthesis in minicells, respectively. The more effective incorporation of 3H-uridine or 14C-leucine into minicells harboring drd7 than ColIb resulted presumably from the high efficiency of the segregation of drd7 into minicells. Polyacrylamide gel electrophoresis of 3H-RNA has shown that plasmids in minicells were able to code low molecular RNA of 4s. No 16 or 23 ribosomal RNA was found in the profiles of de novo synthesized RNA in minicells.