• Journal of Microbiology and Biotechnology
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• v.18 no.2
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• pp.194-198
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• 2008

A newly designed Cytophaga-Flavobacteria-specific 16S rRNA gene primer pair was employed to investigate the CF community structure in the Bering Sea, revealing a previously unknown and unexpected high CF diversity in this high latitude cold sea. In total, 56 clones were sequenced and 50 unique CF 16 rRNA gene fragments were obtained, clustering into 16 CF subgroups, including nine cosmopolitan subgroups, five psychrophilic subgroups, and two putatively autochthonous subgroups. The majority of sequences (82%) were closely related to uncultured CF species and could not be classified into known CF genera, indicating the presence of a large number of so-far uncultivated CF species in the Bering Sea.

• Journal of Microbiology and Biotechnology
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• v.16 no.1
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• pp.77-83
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• 2006

A gene encoding a $\gamma-butyrolactone$ autoregulator receptor was cloned from Saccharopolyspora erythraea, and the biochemical characteristics, including the autoregulator specificity, were determined with the purified recombinant protein. Using primers designed for the conserved amino acid sequence of Streptomyces $\gamma-butyrolactone$ autoregulator receptors, a 120 bp S. erythraea DNA fragment was obtained by PCR. Southern and colony hybridization with the 120 bp fragment as a probe allowed to select a genomic clone of S. erythraea, pESG, harboring a 3.2 kb SacI fragment. Nucleotide sequencing analysis revealed a 615 bp open reading frame (ORF), showing moderate homology (identity, $31-34\%$; similarity, $45-47\%$) with the $\gamma-butyrolactone$ autoregulator receptors from Streptomyces sp., and this ORF was named seaR (Saccharopolyspora erythraea autoregulator receptor). The seaR/pET-3d plasmid was constructed to overexpress the recombinant SeaR protein (rSeaR) in Escherichia coli, and the rSeaR protein was purified to homogeneity by DEAE-Sephacel column chromatography, followed by DEAE-ion-exchange HPLC. The molecular mass of the purified rSeaR protein was 52 kDa by HPLC gel-filtration chromatography and 27 kDa by SDS-polyacrylamide gel electrophoresis, indicating that the rSeaR protein is present as a dimer. A binding assay with tritium-labeled autoregulators revealed that rSeaR has clear binding activity with a VB-C-type autoregulator as the most effective ligand, demonstrating for the first time that the erythromycin producer S. erythraea possesses a gene for the $\gamma-butyrolactone$autoregulator receptor.

• Korean Journal of Microbiology
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• v.51 no.3
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• pp.256-262
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• 2015

In order to study the effect of the receptor protein (SeaR), which is isolated from Saccharopolyspora erythraea, we introduced the SeaR gene to Streptomyces virginiae as host strains. An effective transformation procedure for S. virginiae was established based on transconjugation by Escherichia coli ET12567/pUZ8002 with a ${\varphi}C31$-derived integration vector, pSET152, which contained int, oriT, attP, and $ermEp^{\ast}$ (erythromycin promotor). Therefore, the pEV615 was introduced into S. virginiae by conjugation and integrated at the attB locus in the chromosome of the recipients by the ${\varphi}C31$ integrase (int) function. Transformants of S. virginiae containing the SeaR gene were confirmed by PCR and transcriptional expression of the SeaR gene in the transformants was analyzed by RT-PCR, respectively. And, we examined the production time of virginiamycins in the culture media of both the transformants and the wild type. The production time of virginiamycins in the wild type and transformants was the same. When 100 ng/ml of synthetic $VB-C_6$ was added to the state of 6 or 8 hour cultivation of wild type and transformants, respectively, the virginiamycins production was induced, meaning that the virginiamycins production in the wild type was detected 2 h early than transformants. From these results, SeaR expression was also affected to virginiamycins production in transformants derived from S. virginiae. In this study, we showed that the SeaR protein worked as a repressor in transformants.

• Korean Journal of Microbiology
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• v.51 no.1
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• pp.39-47
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• 2015

Secondary metabolism in actinomycetes has been known to be controlled by a small molecule, ${\gamma}$-butyrolactone autoregulator, the binding of which to each corresponding receptor leads to the regulation of the transcriptional expression of the secondary metabolites. We expected that expression of an autoregulator receptor or a pleiotropic regulator in a non-host was to be gained insight of effective production of new metabolic materials. In order to study the function of the receptor protein (seaR), which is isolated from Saccharopolyspora erythraea, we introduced the seaR gene to Streptomyces coelicolor A3(2) as host strains. An effective transformation procedure for S. coelicolor A3(2) was established based on transconjugation by Escherichia coli ET12567/pUZ8002 with a ${\varphi}C31$-derived integration vector, pSET152, which contained int, oriT, attP and $ermEp^*$ (erythromycin promotor). Therefore, the pEV615 was introduced into S. coelicolor A3(2) by conjugation and integrated at the attB locus in the chromosome of the recipients by the ${\varphi}C31$ integrase (int) function. Exconjugant of S. coelicolor A3(2) containing the seaR gene was confirmed by PCR and transcriptional expression of the seaR gene in the transformant was analyzed by RT-PCR. In case of S. coelicolor A3(2), a phenotype microarray was used to analyze the phenotype of transformant compared with wild type by seaR expression. After that, in order to confirm the accuracy of the results obtained from the phenotype microarray, an antimicrobial susceptibility test was carried out. This test indicated that sensitivity of the transformant was higher than wild type in tetracycline case. These results indicated that some biosynthesis genes or resistance genes for tetracycline biosynthesis in transformant might be repressed by seaR expression. Therefore, subsequent experiments, analysis of transcriptional pattern of genes for tetracycline production or resistance, are needed to confirm whether biosynthesis genes or resistance genes for tetracycline are repressed or not.

• Proceedings of the Korea Society of Environmental Toocicology Conference
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• 2004.11a
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• pp.22-23
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• 2004

• Journal of Microbiology and Biotechnology
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• v.17 no.11
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• pp.1743-1750
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• 2007

Two Gram-negative, nonmotile, coccobacilli, SW-$3^T$ and SW-$100^T$, were isolated from sea water of the Yellow Sea in Korea. Strains SW-$3^T$ and SW-$100^T$ contained ubiquinone-9 (Q-9) as the predominant respiratory lipoquinone and $C_{18:1}\;{\omega}9c$ and $C_{16:0}$ as the major fatty acids. The DNA G+C contents of strains SW-$3^T$ and SW- $100^T$ were 44.1 mol% and 41.9 mol%, respectively. A neighbor-joining tree based on l6S rRNA gene sequences showed that the two isolates fell within the evolutionary radiation enclosed by the genus Acinetobacter. Strains SW-$3^T$ and SW-$100^T$ exhibited a l6S rRNA gene similarity value of 95.7% and a mean DNA-DNA relatedness level of 9.2%. Strain SW-$3^T$ exhibited l6S rRNA gene sequence similarity levels of 93.5-96.9% to the validly described Acinetobacter species and fifteen Acinetobacter genomic species. Strain SW-$100^T$ exhibited l6S rRNA gene sequence similarity levels of less than 97.0% to the other Acinetobacter species except Acinetobacter towneri DSM $14962^T$ (98.0% similarity). Strains SW-$3^T$ and SW-$100^T$ exhibited mean levels of DNA-DNA relatedness of 7.3-l6.7% to the type strains of some phylogenetically related Acinetobacter species. On the basis of phenotypic, phylogenetic, and genetic data, strains SW-$3^T$ and SW-$100^T$ were classified in the genus Acinetobacter as two distinct novel species, for which the names Acinetobacter marinus sp. novo (type strain SW-$3^T$=KCTC $12259^T$=DSM $16312^T$) and Acinetobacter seohaensis sp. novo (type strain SW-$100^T$=KCTC $12260^T$=DSM $16313^T$) are proposed, respectively.

• Journal of Life Science
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• v.15 no.6 s.73
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• pp.879-883
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• 2005

This study was differentiated between Korea and China arkshells using PCR-aided RFLP method which could identify the variation for inter-and intra-species of arkshell (Scapharca broughtonii Schrenck) at the level of DNA. The DNA fragment patterns were compared after digesting gene of mitochondrial 16S rDNA with 8 kinds of restriction enzymes. A 720 bp DNA fragment corresponding to 16S rDNA gene was amplified by PCR with primers ArkF-3 and ArkR-3. PCR products were cut by restriction enzymes (Pvull, BamHI, Hinfl, HaeIII, EcoRI, RsaI, Ksp221, and BstX21), and RFLP pattern was studied. A unique 275 bp DNA band was observed in the samples from Dukyang, Gamak, Namhae, Jinhae, and Taean in Korea when treated by Hinfl, but Chinese arkshell did not show. Treatment of HaeIII could discriminate the sample of Namhae and Jinhae from Dukyang/Gamak/Taean, as well as Korean and Chinese arkshell based on a 700 bp. However, PuvII, BamHI, EcoRI, RsaI, Ksp221, and BstX21 showed the same of 700 bp band in Korean and Chinese arkshell. The phylogenetic tree inferred from PCR-RFLP pattern comparsion in Korean arkshell was different that the distance between Dukyang/Gamak/Taean and Namhae/Jinhae was approximately 7. In particular, the distance between Korean and Chinese arkshell was 25. Consequently, HinfI and HaeIII played an important role in a reliable molecular tool for rapid discriminating Korean and Chinese arkshell, as well as a intra-species in Korea.

• Ocean and Polar Research
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• v.32 no.3
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• pp.309-319
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• 2010

To assess community structure and diversity of archaea, a clone sequencing analysis based on an archaeal 16S rRNA gene was conducted at three sediment depths of the continental slope and Ulleung Basin in the East Sea. A total of 311 and 342 clones were sequenced at the slope and basin sites, respectively. Marine Group I, which is known as the ammonia oxidizers, appeared to predominate in the surface sediment of both sites (97.3% at slope, 88.5% at basin). In the anoxic subsurface sediment of the slope and basin, the predominant archaeal group differed noticeably. Marine Benthic Group B dominated in the subsurface sediment of the slope. Marine Benthic Group D and Miscellaneous Crenarchaeotal Group were the second largest archaeal group at 8-9 cm and 18-19 cm depth, respectively. Marine Benthic Group C of Crenarchaeota occupied the highest proportion by accounting for more than 60% of total clones in the subsurface sediments of the basin site. While archaeal groups that use metal oxide as an electron acceptor were relatively more abundant at the basin sites with manganese (Mn) oxide-enriched surface sediment, archaeal groups related to the sulfur cycle were more abundant in the sulfidogenic sediments of the slope. Overall results indicate that archaeal communities in the Ulleung Basin show clear spatial variation with depth and sites according to geochemical properties the sediment. Archaeal communities also seem to play a significant role in the biogeochemical carbon (C), nitrogen (N), sulfur (S), and metal cycles at each site.

• Journal of Life Science
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• v.19 no.6
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• pp.711-719
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• 2009

We determined a portion of mitochondrial 16S rRNA gene sequences (416 bp) to investigate the genetic structure of the octopus (Octopus minor Ssaki) population in Korea and China. Samples were obtained from Korea (Yeosu, Namhae, Jindo, Muan, Geomundo and Seosan) and China (Sandong) during the period of August 2006 to September 2007. Sequence analyses of 28 individual specimens collected from 7 localities revealed 11 haplotypes, ranging in a sequence divergence of 0.2% - 1.2%. Phylogenetic analyses using PHYLIP and networks subdivided the octopus into two clades (termed clade A and B) and the nucleotide divergence between them was 0.4%. This haplotype subdivision was in accordance with geographic separation: one at Yeosu, Namhae, Muan and Jindo, and the other at Seosan, Geomundo and Sandong. On the basis of hierarchial genetic analysis, genetic distance between localities in Korea and China were also found, but a significant population differentiation was not shown in this study (p>0.05). Consequently, most of the octopus populations in Korea had considerable distribution due to the mitochondrial gene flow that resulted in a formation of a genetically homogenous structure, whereas some of the Korean and Chinese populations had different genetic structures. Gene flow among populations may be restricted due to impassable geographic barriers that promote genetic differentiation.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.14 no.6
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• pp.3114-3119
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• 2013

The mangrove ecosystems have the capacity to act as a sink of heavy metals entering aquatic ecosystems. Despite their potential exposure to metal contaminated sediments, mangroves appear to be highly tolerant to heavy metals. In this study, we cloned metal tolerance gene from mangrove plant. Using CTAB method, RNA were isolated from leaves and root tissue of Rhizophora stylosa habitated at Weno island in Micronesia Chuuk lagoon using CTAB method and phytochelatin synthase 1 (PCS1) gene was cloned using gene specific primers. Expression of PCS1 gene was increased 1.91 fold and 2.72 fold in mangrove propagules exposed to 100 ppb Cd and 10 ppb Cu, respectively. These results indicate that expression of PCS1 gene are promising tools for health assessment of mangrove ecosystem.