Aims: To characterize the transposition mechanism of the IS-element IS10R and study how this element is involved in gene disruption in Lactococcus lactis.
Methods and results: The gene flciA confers immunity against lactococcin A in lactococci. However, the immunity function was lost when flciA was co-expressed with the regulator gene nisR on a plasmid in L. lactis NZ9000. By PCR and DNA sequencing, it was revealed that flciA in immune-negative transformants was disrupted by the IS-element IS10R. Such gene disruption did not occur when flciA was expressed alone nor when the plasmid-located nisR was mutated, suggesting that nisR is directly involved in the transposition. The sequence 5'-CACTTAACC-3', which was found in flciA and at both ends of the inserted IS10R, was identified as target site by site-directed mutagenesis.
Conclusions: IS10R transposes in L. lactis NZ9000 in a nisR-dependent fashion and employs the sequence 5'-CACTTAACC-3' as integration site.
Significance and impact of the study: To our knowledge, this is the first time IS10R and aspects of its transposition are described in the industrial important bacterium L. lactis. The highly controllable insertion of IS10R into a target site might present a great potential as a gene disruption system.