Electroporation of DNA sequences from the pathogenicity locus (PaLoc) of toxigenic Clostridium difficile into a non-toxigenic strain
Abstract
Toxigenic Clostridium difficile is the etiologic agent of C. difficile-associated diarrhoea (CDAD), the most common cause of hospital-acquired infectious diarrhoea. The genes tcdA and tcdB, which encode for the toxin A and B proteins, are part of the pathogenicity locus (PaLoc) of toxigenic C. difficile. Genetic and virulence studies at the molecular level in C. difficile have been hindered by the lack of techniques for DNA manipulation in this species. We describe the electroporation of DNA fragments from a toxigenic isolate into a non-toxigenic strain of C. difficile. Using previously described methods of electroporation into Clostridium spp., the complete toxin B gene and polymerase chain reaction (PCR) fragments of the PaLoc were cloned and electroporated into a non-toxigenic strain of C. difficile. The resulting transformed clones were screened for the introduced gene fragments by PCR, which confirmed their presence. This is the first description of introduction of DNA into C. difficile by electroporation.
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Some well-studied non-acetogenic clostridia, such as C. acetobutylicum, C. beijerinckii, C. difficile, and C. botulinum, have transformation protocols that were established as early as 1988.
“Some of the well-studied, non-acetogenic clostridia, such as the ABE model organisms C. acetobutylicum (Mermelstein et al., 1992) and C. beijerinckii (Oultram et al., 1988), and important pathogens, such as C. difficile (Ackermann et al., 2001) and C. botulinum (Zhou and Johnson, 1993), have transformation protocols that were established as early as 1988.”
From Gas Fermentation—A Flexible Platform for Commercial Scale Production of Low-Carbon-Fuels and Chemicals from Waste and Renewable Feedstocks (2016) · cites this paper as Ackermann et al., 2001
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- Authors
- Ackermann, G., Tang, Y. J., Henderson, J. P., Rodloff, A. C., Silva Jr, J., & Cohen, S. H.
- Affiliations
- University of California Davis Medical Center (US)
- Venue
- Molecular and Cellular Probes
- Publisher
- Elsevier BV
- Published
- 2001-10
- Volume
- 15
- Issue
- 5
- Pages
- 301-306
- ISSN
- 0890-8508 (ISSN-L)
- OpenAlex
- W2143640115
- Cited by
- 11
- References
- 13
- Citations by year
- 2020: 2 · 2016: 1 · 2014: 2 · 2013: 1
- FWCI
- 0.51
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Molecular and Cellular Probes (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Health Sciences › Medicine › Infectious Diseases › Clostridium difficile and Clostridium perfringens research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biotechnology › Microbial Inactivation Methods
- Keywords
- Clostridium difficile, electroporation, pathogenicity locus, toxin B gene, TcdA, TcdB, non-toxigenic strains, toxigenic strains, gene transfer, virulence genes
- MeSH terms
- Bacterial Proteins, Bacterial Toxins, Cloning, Molecular, Clostridium Infections, DNA, Bacterial, Humans, Virulence, Polymerase Chain Reaction, Clostridioides difficile, Electroporation
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