Instability of hybrid plasmids in Bacillus subtilis

We studied the mechanisms responsible for instability of hybrid derivatives of staphylococcal R plasmids in B. subtilis cells. Four pBCH plasmids were constructed from staphylococcal chloramphenicol-resistance plasmid pC756 ligated with pBR322 or pUC8 vectors, and two pAEC plasmids contained pC756 plasmid, erm gene from S. aureus pE3692 and pBR322 plasmid vector. The level of instability of these recombinant plasmids was clearly related to their sizes, the larger derivatives were segregationally highly unstable. The four hybrid pBE plasmids constructed from erythromycin-resistant pE3692 plasmid or its deletion derivative and pBR322 vector were poorly maintained in B. subtilis rec+ strain in comparison with the B. subtilis (recE4) strain. Moreover, the absence in pBE-hybrids of a part of palA sequence led to considerable reduction of plasmid stability in B. subtilis cells. However, the clones of B. subtilis harbouring pBE plasmids integrated into the bacterial chromosome presented much higher stability of the plasmid erm gene than the clones maintaining autonomously replicating plasmids.