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Research Support, Non-U.S. Gov'ts
- A Quantitative and Direct PCR Assay for the Subspecies-Specific Detection of Clavibacter michiganensis subsp. michiganensis Based on a Ferredoxin Reductase Gene
- Min Seok Cho , Jang Ha Lee , Nam Han Her , ChangKug Kim , Young-Joo Seol , Jang Ho Hahn , Ji Hyoun Baeg , Hong Gi Kim , Dong Suk Park
- J. Microbiol. 2012;50(3):496-501. Published online June 30, 2012
- DOI: https://doi.org/10.1007/s12275-012-1611-x
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- 3 Scopus
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Abstract
- The Gram-positive bacterium Clavibacter michiganensis subsp. michiganensis is the causal agent of canker disease in tomato. Because it is very important to control newly introduced inoculum sources from commercial materials, the specific detection of this pathogen in seeds and seedlings is essential for effective disease control. In this study, a novel and efficient assay for the detection and quantitation of C. michiganensis subsp. michiganensis in symptomless tomato and red pepper seeds was developed. A pair of polymerase chain reaction (PCR) primers (Cmm141F/R) was designed to amplify a specific 141 bp fragment on the basis of a ferredoxin reductase gene of C. michiganensis subsp. michiganensis NCPPB 382. The specificity of the primer set was evaluated using purified DNA from 16 isolates of five C. michiganensis subspecies, one other Clavibacter species, and 17 other reference bacteria. The primer set amplified a single band of expected size from the genomic DNA obtained from the C. michiganensis subsp. michiganensis strains but not from the other C. michiganensis subspecies or from other Clavibacter species. The detection limit was a single cloned copy of the ferredoxin reductase gene of C. michiganensis subsp. michiganensis. In conclusion, this quantitative direct PCR assay can be applied as a practical diagnostic method for epidemiological research and the sanitary management of seeds and seedlings with a low level or latent infection of C. michiganensis subsp. michiganensis.
- The Involvement of the nif-Associated Ferredoxin-Like Genes fdxA and fdxN of Herbaspirillum seropedicae in Nitrogen Fixation
- André L.F. Souza , Adriana L. Invitti , Fabiane G.M. Rego , Rose A. Monteiro , Giseli Klassen , Emanuel M. Souza , Leda S. Chubatsu , Fábio O. Pedrosa , Liu U. Rigo
- J. Microbiol. 2010;48(1):77-83. Published online March 11, 2010
- DOI: https://doi.org/10.1007/s12275-009-0077-y
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- 9 Scopus
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Abstract
- The pathway of electron transport to nitrogenase in the endophytic β-Proteobacterium Herbaspirillum seropedicae has not been characterized. We have generated mutants in two nif-associated genes encoding putative ferredoxins, fdxA and fdxN. The fdxA gene is part of the operon nifHDKENXorf1orf2fdxAnifQmodABC and is transcribed from the nifH promoter, as revealed by lacZ gene fusion. The fdxN gene is probably cotranscribed with the nifB gene. Mutational analysis suggests that the FdxA protein is essential for maximum nitrogenase activity, since the nitrogenase activity of the fdxA mutant strain was reduced to about 30% of that of the wild-type strain. In addition, the fdxA mutation had no effect on the nitrogenase switch-off in response to ammonium. Nitrogenase activity of a mutant strain lacking the fdxN gene was completely abolished. This phenotype was reverted by complementation with fdxN expressed under lacZ promoter control. The results suggest that the products of both the fdxA and fdxN genes are probably involved in electron transfer during nitrogen fixation.
- Association of a Common Reductase with Multiple Aromatic Terminal Dioxygenases in Sphingomonas yanoikuyae Strain B1
- Mihyun Bae , Eungbin Kim
- J. Microbiol. 2000;38(1):40-43.
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Abstract
- The aromatic dioxygenase system in Sphingomonas yanoikuyae strain B1 consists of three components, an oxygenase, a ferredoxin, and a reductase. The insertional knockout of the bphA4 gene encoding a reductase and subsequent complementation experiments showed that the reductase encoded by bphA4 in S. yanoikuyae strain B1 is associated with multiple dioxygenase components including that of toluate dioxygenase (XylXY).
- Molecular Cloning and Analysis of the Gene for P-450 Hydroxylase from Pseudonocardia autotrophica IFO 12743
- Jung-Mee Kim , Younmie Jin , Chang-Gu Hyun , Jong-Hee Kim , Hong-Sub Lee , Dae-Kyung Kang , Dae-Jung Kang , Tae-Yong Kim , Joo-Won
- J. Microbiol. 2002;40(3):211-218.
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Abstract
- A 4.8-kb DNA fragment encoding the P-450 type hydroxylase and ferredoxin genes was cloned from Pseudonocardia autotrophica IFO 12743 that can convert vitamin D_3 into its hydroxylated active forms. In order to isolate the P-450 gene cluster in this organism, we designed PCR primers on the basis of the regions of an oxygen binding site and a heme ligand pocket that are general characteristics of the P-450 hydroxylase. Sequencing analysis of the BamHI fragment revealed the presence of four complete and one incomplete ORFs, named PauA, PauB, PauC, and PauD, respectively. As a result of computer-based analyses, PauA and PauB have homology with enoyl-CoA hydratase from several organisms and the positive regulators belonging to the tetR family, respectively. PauC and PauD show similarity with SuaB/C proteins and ferredoxins, respectively, which are composed of P-450 monooxygenase systems for metabolizing two sulfonylurea herbicides in Streptomyces griseolus PauC shows the highest similarity with another CytP-450_Sca2 protein that is responsible for production of a specific HMG-CoA reductase inhibitor, pravastatin, in S. carbophilus. Cultures of Streptomyces lividans transformant, containing the P-450 gene cluster on the pWHM3 plasmid, was unable to convert vitamin D_3 to its hydroxylated forms.
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