| Project/Area Number |
09557024
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 展開研究 |
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| Research Field |
Bacteriology (including Mycology)
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| Research Institution | GIFU UNIVERSITY |
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Principal Investigator |
EZAKI Takayuki GIFU UNIV.MICROBIOLOGY,PROFESSOR, 医学部, 教授 (90151977)
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| Co-Investigator(Kenkyū-buntansha) |
KAWAMURA Yoshiaki GIFU UNIV.MICROBIOLOGY,ASSISTANT, 医学部, 助手 (80262757)
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| Project Period (FY) |
1997 – 1998
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| Project Status |
Completed (Fiscal Year 1998)
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| Budget Amount *help |
¥9,500,000 (Direct Cost: ¥9,500,000)
Fiscal Year 1998: ¥3,800,000 (Direct Cost: ¥3,800,000)
Fiscal Year 1997: ¥5,700,000 (Direct Cost: ¥5,700,000)
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| Keywords | taxonomy / species definition / DNA hybrid / ribosome / Phylogeny / Liquid phase hybridization / リボソーム / 16S リボゾーム / DNA 相同性 / 種 |
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| Research Abstract |
Prokaryotes were phylogenetically classified by 16SrRNA sequences and genetic distance among independent species were quantitatively measured simply by comparing their 16SrRNA differences. However, it become clear that some established species shares same 16SrRNA sequences.
In this case, we need to use other method to differentiate closely related species. Quantitative DNA/DNA hybridization method is a officially accepted method to define a bacterial species. However, this method is difficult to perform and DNA used for this method is often cotaminated with polysaccharide, which interfers quantitative analysis of DNA/DNA hybrid. We established several parameters to evaluate DNA purity and a rapid measuring method of guanine plus cytosine contents of DNA to define DNA/DNA hybridization experiments. RNA and polysaccharide contamination was successfully predicted by measuring absorbance of 235/260 nm and Anthrone quantitation method of sugars. Guanine plus cytosine content of DNA was instantly measured by Light cycler with SYBR Green 1 fluorescence. This information is very useful to define quantitative DNA/DNA hybridization at optimal temperature. Trials to differentiate strains within a species were successfully introduced by many scientists. We used Ceul restriction enzyme to evaluate distances among strains. This method has a advantage because bands generated by the Ceul digestion indicate numbers of copies of ribosomal RNA operon. From fragment analysis of generated size offered useful information to analyze gene recombination history.
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