2 μm plasmid copy number in different yeast strains and repartition of endogenous and 2 μm chimeric plasmids in...

By using the renaturation kinetics technique we tried to get informations about the maintenance of the 2 μm plasmid in yeast cells. For this purpose we determined the 2 μm plasmid copy number: in various yeast strains, in a special set of mutants, in cells treated with ethidium bromide and cycloheximide and in different yeast strains obtained by transformation with 2 μm chimeric plasmids.According to the strain used the proportion of 2μm DNA varied from 1.1% to 3.9%, which corresponds to 24 to 88 2 μm molecules per haploid genome. The particular multiresistant mutant, where the frequent loss of oligomycine resistance is correlated with the loss of extractible covalently closed circular DNA, contained 39 2 μm copies per haploid genome. In the partial revertant oligomycine sensitive all the 2 μm DNA sequences were lost. (Less than 0.1 copy per haploid genome.)Ethidium bromide did not affect the 2 μm copy number while cycloheximide induces an increase of 36%.When a strain containing 88 2 μm DNA copies per haploid genome is transformed with 2 μm chimeric plasmids there is no significative change in the total number of plasmid: 36 copies of endogenous and 44 of chimeric plasmid per haploid genome. When 2 μm chimeric plasmids were introduced in our 2 μm-less strain despite the stability of the transformants, there is only 8 copies per haploid genome.