2006 Fiscal Year Final Research Report Summary
Isolation of dechlorinating genes and enzymes from soil of the long-term experimental field where organochlorine pesticides have been applied for more than thirty years
| Project/Area Number |
16380047
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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 | 一般 |
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| Research Field |
Plant nutrition/Soil science
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| Research Institution | The University of Tokyo |
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Principal Investigator |
SENOO Keishi The University of Tokyo, Graduate School of Agricultural and Life Sciences, Professor (40206652)
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| Co-Investigator(Kenkyū-buntansha) |
OTSUKA Shigeto The University of Tokyo, Graduate School of Agricultural and Life Sciences, Lecturer (10313074)
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| Project Period (FY) |
2004 – 2006
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| Keywords | soil / long-term experimental field / γ-HCH / soil DNA / lin genes / dioxygenase genes / soil microoraanisms / ecosystem restoration |
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| Research Abstract |
1. Isolation of IinB (haloalkane dehalogenase) with sequence diversity
Five strains of novel γ-HCH-degrading Sphingbium were obtained without enrichment culture. PCR amplification of genomic DNA of these strains targeting lin genes followed by cloning and sequencing were carried out. As a result, five different linB with nucleotide sequence diversity were obtained.
PCR amplification of soil DNA extracted from γ-HCH-added manure plot soil targeting genes flanked by IS6100 was carried out. Cloning and sequencing of the amplified product revealed that all of the analysed /IS6100-flanked genes were linB with sequence diversity.
2. Isolation of linA (dehaydrochlorinase) with sequence diversity
linA-targeting PCR amplification of soil DNA extracted from different plots in the long-term experimental field was carried out. The amplified products were obtained for all of the soil DNA. Cloning and sequencing of the DNA generated several linA-homologous DNA sequences with several base substitutions.
3. Isolation of petroleum-degrading genes with sequence diversity
Soils collected from different plots (manure, TPN/manure, TPN/HCH/manure) in the long-term experimental field were added with petroleum component(naphthalene, fluorine and petroleum ether) and incubated. Soil DNA extracted from the incubated soils were subjected to PCR trying to amplify naphthalene degrading genes. After cloning and sequencing analysis of the amplified products, dioxygenase gene-like DNA sequences were obtained. Not only DNA sequences highly homologous to that of known dioxygenase, but also a number of novel sequences with low homology were obtained.
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