Bacitracins are a mixture of structurally related cyclic polypeptides with clinically useful antibiotic properties. They act by indirectly interfering with the biosynthesis of the bacterial cell wall. In this study, we analysed an avian necrotic enteritis strain of Clostridium perfringens that was resistant to bacitracin. We identified, for the first time in this genus, a putative bacitracin resistance locus that resembles a bacitracin resistance determinant from Enterococcus faecalis. It contains the structural genes bcrABD and a putative regulatory gene, bcrR. Conjugation experiments provided evidence that this locus was located on a mobile genetic element that had the ability to transpose onto both conjugative and non-conjugative plasmids or onto the chromosome. Whole genome high-throughput sequencing of the wild-type strain and several transconjugants revealed that the bcr genes were located on a potential mobile element that was related to the Tn916 family of conjugative transposons. Our analysis identified the open reading frames (ORFs) that are responsible for recombination (Xis/Int), regulation (ORF8), conjugation (ORFs13-23) and several hypothetical genes. The bacitracin resistance locus was located between genes that were homologous to ORF19 and ORF18 from Tn916. In summary, we have identified a novel mobile bacitracin resistance determinant in a toxin-producing C. perfringens strain and our study has provided important bioinformatic and functional information on the genetic organization of this resistance locus.