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We designed a novel DNA probe and novel PCR primer sets to detect the genes coding for S. hyicus exfoliative toxin (ET). In dot blot hybridization, the novel DNA probe hybridized with chromosomal DNA of ExhA-, ExhB-, ExhC-, ExhD-, and SHETA-producing strains. This probe also hybridized with plasmid DNA of a SHETB-producing strain. In Southern blot hybridization, the probe hybridized with a 1.5-kb HindIII fragment of chromosomal DNA from a SHETA-producing strain. We cloned the above fragment into E. coli and determined the nucleotide sequence of the SHETA gene, which proved to be almost the same (99.6% homology) as that of the ExhB gene. So it was thought that SHETA is a subtype of ExhB. In multiplex PCR using five primer sets, each gene gave a band distinguishable from the others. This multiplex PCR system had high specificity among the well-known S. hyicus ET genes. Of the 69 known ET-producing S. hyicus strains, 38, 19, 9, 2 and 1 strains have exhB, exhD exhA, shetb and exhC genes, respectively.
Laboratory of Veterinary Microbiology, Faculty of Veterinary Medicine, School of Veterinary Medicine, Kitasato University, 35-1, Higashi 23 Ban-cho, Towada, 034-8628 Aomori, Japan.
This article was published in the following journal.
Name: Microbiology and immunology
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Species- or subspecies-specific DNA (including COMPLEMENTARY DNA; conserved genes, whole chromosomes, or whole genomes) used in hybridization studies in order to identify microorganisms, to measure DNA-DNA homologies, to group subspecies, etc. The DNA probe hybridizes with a specific mRNA, if present. Conventional techniques used for testing for the hybridization product include dot blot assays, Southern blot assays, and DNA:RNA hybrid-specific antibody tests. Conventional labels for the DNA probe include the radioisotope labels 32P and 125I and the chemical label biotin. The use of DNA probes provides a specific, sensitive, rapid, and inexpensive replacement for cell culture techniques for diagnosing infections.
Synthetic or natural oligonucleotides used in hybridization studies in order to identify and study specific nucleic acid fragments, e.g., DNA segments near or within a specific gene locus or gene. The probe hybridizes with a specific mRNA, if present. Conventional techniques used for testing for the hybridization product include dot blot assays, Southern blot assays, and DNA:RNA hybrid-specific antibody tests. Conventional labels for the probe include the radioisotope labels 32P and 125I and the chemical label biotin.
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