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Genotypic Differentiation of Twelve Clostridium Species by Polymorphism Analysis of the Triosephosphate Isomerase (tpi) Gene
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Genotypic Differentiation of Twelve Clostridium Species by Polymorphism Analysis of the Triosephosphate Isomerase (tpi) Gene
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Genotypic Differentiation of Twelve Clostridium Species by Polymorphism Analysis of the Triosephosphate Isomerase (tpi) Gene
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Journal Article
Genotypic Differentiation of Twelve Clostridium Species by Polymorphism Analysis of the Triosephosphate Isomerase (tpi) Gene
2003
Overview
Housekeeping genes encoding metabolic enzymes may provide alternative markers to 16S ribosomal DNA (rDNA) for genotypic and phylogenetic characterization of bacterial species. We have developed a PCR-restriction fragment length polymorphism (PCR-RFLP) assay, targeting the triosephosphate isomerase (
tpi) gene, which allows the differentiation of twelve pathogenic
Clostridium species. Degenerate primers constructed from alignments of
tpi sequences of various Gram-positive bacteria allowed the amplification of a 501 bp target region in the twelve
Clostridium type strains. A phylogenetic tree constructed from the nucleotidic sequences of these
tpi amplicons was well correlated with that inferred from analysis of 16S rDNA gene sequences. The analysis of
tpi sequences revealed restriction sites of enzyme
Alu I that could be species-specific. Indeed,
Alu I digestion of amplicons from the twelve type strains provided distinct restriction patterns. A total of 127 strains (three to sixteen strains for each species) was further analyzed by PCR-RFLP of the
tpi gene, and confirmed that each species could be characterized by one to three restriction types (RTs). The differences between RTs within species could be explained by point mutations in
Alu I restriction sites of the
tpi sequences. PCR-restriction analysis of the
tpi gene offers an accurate tool for species identification within the genus
Clostridium, and provides an alternative marker to 16S rDNA for phylogenetic analyses.
Publisher
Elsevier GmbH,Elsevier,Elsevier Science Ltd
Subject
/ Biological and medical sciences
/ Clostridium - classification
/ Clostridium - isolation & purification
/ Clostridium– genotypic differentiation – tpi housekeeping gene
/ Electrophoresis, Agar Gel - methods
/ Fundamental and applied biological sciences. Psychology
/ Growth, nutrition, metabolism, transports, enzymes. Molecular biology
/ Microbiology and Parasitology
/ Mycology
/ Polymorphism, Restriction Fragment Length
/ Replicative cycle, interference, host-virus relations, pathogenicity, miscellaneous strains
/ RNA, Ribosomal, 16S - analysis
/ Triose-Phosphate Isomerase - analysis
/ Triose-Phosphate Isomerase - genetics
/ Virology
