Analysis of genes concerned with bacterial magnetite production
| Project/Area Number |
03453128
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
発酵工学
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| Research Institution | Tokyo University of Agriculture & Technology |
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Principal Investigator |
MATSUNAGA Tadashi Tokyo University of Agriculture & Technology Department of Biotechnology, Professor, 工学部, 教授 (10134834)
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| Co-Investigator(Kenkyū-buntansha) |
NAKAMURA Noriyuki Tokyo University of Agriculture & Technology Department of Biotechnology, Assist, 工学部, 助手 (20198229)
SODE Koji Tokyo University of Agriculture & Technology Department of Biotechnology, Associ, 工学部, 助教授 (10187883)
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| Project Period (FY) |
1991 – 1992
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| Project Status |
Completed (Fiscal Year 1992)
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| Budget Amount *help |
¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1992: ¥1,900,000 (Direct Cost: ¥1,900,000)
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| Keywords | Magnetic bacterium / Transposon mutant / Conjugation / Magnetic particles / Iron transport / 16S rRNA / MagA gene / 磁性細菌粒子 / 鉄還元 |
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| Research Abstract |
1. Development of a gene transfer system for magnetic bacteria.
Broad-host-range plasmids have been transferred to the aerobic magnetic bacterium Magnetospirillum sp. AMB-1. Conjugal matings with Escherichia coli S17-1 allowed high frequency transfer of the RK2 derivative pRK415 (4.5 x 10^<-3> transconjugants per recipient cell) and the RSF-1010 derivative pKT230 (3.0 x 10^<-3>). Optimum mating time was 6 hrs for both plasmids. These plasmids successfully formed autonomous replicons in transconjugants and could be isolated and transformed back into E.coli, illustrating their potential as shuttle vectors.
2. Genetic analysis of transposon mutant, NM5.
The non-magnetic mutant NM5 is one of transposon mutagenized mutants. We cloned EcoRI genomic DNA fragment containing the Tn5 interrupted locus. Gene expression and nucleotide sequence of this cloned fragment were analyzed. Dot blot northern hybridization was performed using DNA fragments flanking the Tn5 insertion site, as a probe. Iron suff
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icient and iron limited media were prepared for cultivation of wild type AMB-1 cells from which total RNA was extracted. Strong hybridization was detected for samples from cells cultured in iron limited media. This result suggests that gene(s) whose expression was regulated by iron are present on the cloned EcoRI fragment from NM5. A 2640bp genomic DNA fragment from mutant NM5 was sequenced completely. Three open reading frames were found, one of which is interrupted by the Tn5 insertion. A putative promoter and a ribosome binding site exist upstream of this ORF. This ORF, named magA, was analyzed using the DNA / protein computer analysis software, DNASIS. The predicted amino acid sequence of the magA gene product has high homology with cation transport channel proteins, in particular the E.coli Kef C protein which functions as a potassium efflux protein for control of turgor. Furthermore the amino acid sequence of MagA shows high hydrophobicity. Therefore MagA is probably localized in the membrane fraction and may function as a cation transporter protein in the magnetic bacterium Magnetospirillum sp. AMB-1. Less
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Research Products
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