HR: 0800h
AN: V41B-1372    [Abstracts]
TI: Improved Method for Direct Detection of Environmental Microorganisms Using an Amplification of 16S rDNA Region
AU: * Tsujimura, M
EM: masa.tsujimura@aist.go.jp
AF: Precision System Science Co., Ltd, 88 Kamihongou, Matsudo, 271-0064 Japan
AU: * Tsujimura, M
EM: masa.tsujimura@aist.go.jp
AF: Institute for Biological Resources and Function, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba central 6, 1-1-1 Higashi, Tsukuba, 305-8566 Japan
AU: Akutsu, J
AF: Institute for Biological Resources and Function, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba central 6, 1-1-1 Higashi, Tsukuba, 305-8566 Japan
AU: Zhang, Z
AF: Precision System Science Co., Ltd, 88 Kamihongou, Matsudo, 271-0064 Japan
AU: Zhang, Z
AF: Institute for Biological Resources and Function, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba central 6, 1-1-1 Higashi, Tsukuba, 305-8566 Japan
AU: Sasaki, M
AF: Precision System Science Co., Ltd, 88 Kamihongou, Matsudo, 271-0064 Japan
AU: Sasaki, M
AF: Institute for Biological Resources and Function, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba central 6, 1-1-1 Higashi, Tsukuba, 305-8566 Japan
AU: Tajima, H
AF: Precision System Science Co., Ltd, 88 Kamihongou, Matsudo, 271-0064 Japan
AU: Kawarabayasi, Y
EM: kawarabayasi.yutaka@aist.go.jp
AF: Institute for Biological Resources and Function, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba central 6, 1-1-1 Higashi, Tsukuba, 305-8566 Japan
AB: The thermostable proteins or enzymes were expected to be capable to be utilized in many areas of industries. Many thermophilic microorganisms, which possess the thermostable proteins or enzymes, were identified from the extreme environment. However, many unidentified and uncultivable microorganisms are still remaining in the environment on the earth. It is generally said that the cultivable microorganisms are less than 1% of entire microorganisms living in the earth, remaining over 99% are still uncultivable. As an approach to the uncultivable microorganisms, the PCR amplification of 16S rDNA region using primer sets designed from the conserved region has been generally utilized for detection and community analysis of microorganism in the environment. However, the facts, that PCR amplification introduces the mutation in the amplified DNA fragment and efficiency of PCR amplification is depend on the sequences of primer sets, indicated that the improving of PCR analysis was necessary for more correct detection of microorganisms. As the result of evaluation for the quality of DNA polymerases, sequences of primers used for amplification and conditions of PCR amplification, the DNA polymerase, the primer set and the conditions for amplification, which did not amplify the DNA fragment from the DNA contaminated within the DNA polymerase itself, were successfully selected. Also the rate of mutation in the DNA fragment amplified was evaluated using this conditions and the genomic DNA from cultivable microbes as a template. The result indicated the rate of mutation introduced by PCR was approximately 0.1% to 0.125%. The improved method using these conditions and error rate calculated was applied for the analysis of microorganisms in the geothermal environment. The result indicated that four kinds of dominant microorganisms, including both of bacteria and archaea, were alive within soil in the hot spring in Tohoku Area. We would like to apply this improved method to detection of microorganisms with important genes from more other environments.
DE: 8400 VOLCANOLOGY
DE: 8494 Instruments and techniques
DE: 8499 General or miscellaneous
SC: Volcanology, Geochemistry, Petrology [V]
MN: 2004 AGU Fall Meeting