Integration and excision of plasmid in Bacillus subtilis

We have studied the behaviour in Bacillus subtilis of a plasmid (pPV21) carrying the thymidylate synthetase gene of phage phi3T (thyP3). The plasmid can transform efficiently the competent cells of all the strains tested. Polyethylene glycol (PEG)-mediated protoplast transformation is efficient only for recE, recD or recF mutants. When present in recombination proficient strains, the plasmid can be integrated into the chromosome, primarily at the thyA locus. This has been shown by genetic mapping and by blot-hybridization. A second less efficient site is at (or near to) the attachment site of phage phi3T. Excision of the plasmid restores the EcoRI restriction pattern of the parental DNA, although with the loss of the defective thyA endogenotic allele and the retention of the thyP exogenotic gene.