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Detection of Staphylococcus hyicus exfoliative toxin genes by dot blot hybridization and multiplex polymerase chain reaction
Author(s) -
Onuma Kenta,
Uoya Yusuke,
Koide Tetsuo,
Shibata Ayumi,
Tanabe Taishi,
Sato Hisaaki
Publication year - 2011
Publication title -
microbiology and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.664
H-Index - 70
eISSN - 1348-0421
pISSN - 0385-5600
DOI - 10.1111/j.1348-0421.2011.00308.x
Subject(s) - biology , microbiology and biotechnology , primer (cosmetics) , southern blot , dot blot , gene , multiplex polymerase chain reaction , hybridization probe , polymerase chain reaction , plasmid , genetics , chemistry , organic chemistry
We designed a novel DNA probe and novel PCR primer sets for detecting the genes coding for Staphylococcus hyicus ( 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 the plasmid DNA of a SHETB‐producing strain. In Southern blot hybridization, the probe hybridized with a 1.5 kb Hin dIII fragment of chromosomal DNA from a SHETA‐producing strain. The above fragment was cloned into E. coli and the nucleotide sequence of the SHETA gene determined, this gene proved to have almost the same homology (99.6%) as the ExhB gene. It was therefore 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 has high specificity among the well‐known S. hyicus ET genes. Of the 69 known ET‐producing S. hyicus strains, 38, 19, 10, 2 and 1 strains have exh B, exh D exhA , shetb and exhC genes, respectively.