TcdC consists of three domains: a hydrophobic domain, a proposed dimerization domain and a proposed C-terminal repressor domain ( Figure 1A ) [12] . We successfully disrupted the tcdC gene in the region coding for the repressor domain using ClosTron technology. Disruption of genes using the ClosTron technology results in stable mutants and no or non-functional proteins [24] [25] [26] . The genotype of the disruption was confirmed with conventional PCRs using the tcdC2 primer and the EBS universal primer and with a primer pair (tcdC1 and tcdC2) flanking the ClosTron insertion site ( Figure B ). Sequence analysis confirmed that the disruption was in the proposed repressor domain of the tcdC gene at the expected site (data not shown). In addition, Southern blot analysis using intron-, ermB and tcdCspecific probes clearly confirmed a specific single insertion of the Group II intron in the genome ( Figure 1C ).