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KMID : 0355519920160010045
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Korean Journal of Oral Biology
1992 Volume.16 No. 1 p.45 ~ p.50
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Whole-genomic DNA Probes for the-Identification of Oral Bacteroides Species
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Choe Se-Won
Choe Seon-Jin
Lee Jeong-Sook
Kim Kack-Kyun
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Abstract
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The need for a rapid and sensitive microbiological assay has become necessary for both research and clinical diagnostic purposes. This need has become clear as result of extensive documentation linking specific bacterial species and periodontal destruction. DNA probe technology seems to be able to provide both a sensitive and specific assay and may be able to alleviate the concern for transport of fastidious microorganisms. We have been trying to establish the method of identifying the periodontopathic microorganisms by DNA probe. As the first step we tried to know if whole-genomic DNA probe could differentiate between bacterial species, using three different serotype-strains of both Bacteroides gingivalis and Bacteroides intermedius. Also we tested if there is any cross-hybridization between Bacteroides spp, and streptococcus spp. The experimental procedures were as follows: anaerobic culture of bacterial cells, preparation of whole-genomic DNA from bacteria and measurement of DNA concentration, immobilization of DNA onto a nitrocellulose filter using dot-blot apparatus after. 2-fold serial dilution of DNA. Radio-labelling of DNA with 32P by random oligonucleotide-primed synthesis, which would serve as a DA probe. Blocking ubound nitrocellulose with 0.05×BLOTTO, hybridization of the filter with 32P-labelled probe, and washing and detection of bound probe by autoradiography. The results were as follows. The whole-genomic DNA probe of B. gingivalis strain 381 (serotype a) was able to hybridize with all three serotypes a, b, and c) of the species. Also, the whole-genomic DNA probe of B. intermedius strain 9336 (serotype B) was able to hybridize with all three serotypes of the species. There was no detectable cross-hybridization between B. gingivalis and B. intermedius or bacteroides DNA probe with streptococcal DNA, within the range of DNA amount used in the experiment. The DNA probe was able to detect 3ng of homologous DNA in a dot. It seems likely that whole-genomic probe could be used for identification of Bacteroides spp. in the clinical samples using DNA probe, if a suitable method to lyse the cell on the filter is provided.
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KEYWORD
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DNA probes, periodontal disease, Bacteroides spp, diagnosis
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