Monday, May 3, 2010

Transposon mutagenesis identifies a novel toxin regulatory locus in Clostridium perfringens

Over the years, we’ve had many inquiries about using the EZ-Tn5™ Transposomes on biologically interesting but difficult-to-mutate bacteria--usually Gram-positive bacteria that have poorly understood genetics and are difficult to transform with foreign DNA. As time has progressed, some of the difficulties of using EZ-Tn5 Transposomes have been overcome. An example of such success was recently reported by Vidal et al.* regarding the use of a custom EZ-Tn5 Transposome that confers tetracycline resistance to its targeted host cell. Clostridium perfringens is a Gram-positive, obligate spore-forming anaerobe. Its growth characteristics pose considerable obstacles to the use of transposon mutagenesis in the search for genes associated with virulence factors.

The authors used the pMOD-2 Transposon Construction Vector to build an erythromycin-resistance transposon using the erm gene from the E. coli-C. perfringens shuttle vector pJIR751. The Transposome complex was built using standard procedures and transformed using a Biorad Gene Pulser™ electroporator. Using the EZ-Tn5 system, the researchers were able to locate a mutant that disabled the toxin-13 regulatory locus (determined to be similar to the Agr locus of Staphylococcus aureus). They also reported that the generation of mutant Clostridia using the EZ-Tn5 Transposomes was much more efficient than other random mutagenesis methods.

If you're considering the EZ-Tn5 system for bacterial mutagenesis, visit the EZ-Tn5 Transposomes citation page for information on your species of interest.*Vidal, J. et al. (2009). Use of an EZ-Tn5-Based Random Mutagenesis System to Identify a Novel Toxin Regulatory Locus in Clostridium perfringens Strain 13 PLoS ONE, 4 (7) DOI: 10.1371/journal.pone.0006232

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