Development of an intermolecular transposition assay system in Bacillus subtilis 168 using IS4Bsu1 from Bacillus subtilis (natto).

Most of the spontaneous poly-gamma-glutamate (gamma-PGA)-deficient mutants of Bacillus subtilis (natto) appear to have resulted from the insertion of IS4Bsu1 exclusively into the comP gene. However, complete genomic analysis of B. subtilis 168, a close relative of B. subtilis (natto), revealed no IS4Bsu1 insertion. Preliminary experiments using a transformable 'natto' strain indicated that the frequency of transposition of IS4Bsu1 was exceptionally high under competence-developing conditions. On the other hand, such high-frequency transposition was not observed when cells were grown in a rich medium, such as LB medium, suggesting that there must be suitable environmental conditions that give rise to the transposition of IS4Bsu1. To assess the behaviour of IS4Bsu1 and explore any host factors playing roles in IS transposition, an intermolecular transposition assay system was constructed using a modified IS4Bsu1 element in B. subtilis 168. Here, the details of the intermolecular transposition assay system are given, and the increase in transposition frequency observed under high-temperature and competence-inducing conditions is described.