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. 2004 Jan;42(1):179-85.
doi: 10.1128/JCM.42.1.179-185.2004.

Detection of the Bacillus anthracis gyrA gene by using a minor groove binder probe

William Hurtle  1 , Elizabeth Bode, David A Kulesh, Rebecca Susan Kaplan, Jeff Garrison, Deanna Bridge, Michelle House, Melissa S Frye, Bonnie Loveless, David Norwood
Affiliations

Detection of the Bacillus anthracis gyrA gene by using a minor groove binder probe

William Hurtle et al. J Clin Microbiol. 2004 Jan.

Abstract

Identification of chromosomal markers for rapid detection of Bacillus anthracis is difficult because significant chromosomal homology exists among B. anthracis, Bacillus cereus, and Bacillus thuringiensis. We evaluated the bacterial gyrA gene as a potential chromosomal marker for B. anthracis. A real-time PCR assay was developed for the detection of B. anthracis. After analysis of the unique nucleotide sequence of the B. anthracis gyrA gene, a fluorescent 3' minor groove binding probe was tested with 171 organisms from 29 genera of bacteria, including 102 Bacillus strains. The assay was found to be specific for all 43 strains of B. anthracis tested. In addition, a test panel of 105 samples was analyzed to evaluate the potential diagnostic capability of the assay. The assay showed 100% specificity, demonstrating the usefulness of the gyrA gene as a specific chromosomal marker for B. anthracis.

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Figures

FIG. 1.
FIG. 1.
Nucleotide positions of primers BAGYRA1614F and BAGYRA1732R and probe BAGYRA1668MGB.
FIG. 2.
FIG. 2.
Results of triplicate analysis of B. anthracis Ames 10-fold serial dilutions ranging from 1 ng to 1 fg of template DNA. The average cycle threshold values were as follows: 1 ng, 22.25; 100 pg, 25.16; 10 pg, 28.29; 1 pg, 31.77; and 100 fg, 35.26. The 10-fg dilution was not detected consistently. The 1-fg dilution and the no-template control were not detected.
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