• Journal of Life Science
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• v.32 no.3
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• pp.202-209
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• 2022

The green alga Chlamydomonas reinhardtii, a unicellular haploid eukaryote, has long been used by researchers and industries as a cell factory to produce high value-added microalgae substances using genetic modification. Microalga K01, presumed to be Chlamydomonas, was isolated from 12 freshwater samples from the Chungcheong and Jeolla regions to replace C. reinhardtii, an introduced species currently used in most basic and industrial research. The isolated K01 strain was identified as C. reinhardtii through morphological and phylogenetic studies of the 18S rDNA gene sequence (NCBI accession number KC166137). The growth and lipid content of the isolated C. reinhardtii K01 were compared with three wild and four mutant strains in TAP medium, and it was found that the K01 strain could produce 1.74×10⁷ cells/ml by the third day of culture. The growth rate of C. reinhardtii K01 was 1.5 times faster than UTEX2244, which showed the highest number of cells (1.20×10⁷ cells/ml) among the compared strains. The lipid content of the isolated C. reinhardtii K01 (20.67%) was similar to those of the wild strains, although the fatty acid oleate C18:1 was not detected in the isolated strain but was identified in the seven others. The cell density of the isolated strain increased to 0.87 g/l during a six-day culture in BG11 medium, where nitrate (NaNO₃) was introduced as a nitrogen source, while the seven acquired strains showed almost no cell proliferation.

• Applied Biological Chemistry
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• v.13 no.2
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• pp.153-170
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• 1970

1. Isolation and identification of amylase-producing bacteria. The powerful strain A-12 and S-8 were respectively isolated from air and soil after screening a large number of amylase-producing bacteria. Their bacterial characteristics have been investigated and it has been found that all characteristics of strain A-12 and S-8 are similar to Bac. subtilis of Bergey's manual except for the acid formation from a few carbohydrates and the citrate utilization, i.e., the strain A-12 shows negative in the citrate utilization, and the acid formation from arabinose and xylose, S-8 shows negative in the acid formation from xylose. 2. Amylase production by Liquid cultures with solid materials. Several conditions for amylase production by strain A-12 in stationary cultures have been studied. The results obtained are as follows. (1) The optimum conditions are:temperature $35^{\circ}C$, initial pH 6.5 to 7.0 and incubation time 3 to 4 days. (2) The amylase production is not affected by the preservation period of the stock cultures. (3) Among the various solid material, the defatted soy bean is found to be the best for t1e amylase production. However, the alkali treatment of the defatted soy bean gives no effect contrary to the cage of defatted rape seed. The addition of soluble starch to the alkali extract of defatted soy bean shows the increased amylase production. (4) Up to 1% addition of ethanol to carbon dificient media gives the improved amylase production, whereas the above effect is not found in the case of carbon rich media. (5) The amylase production can be increased 2.5 times when 10% of defatted soy bean is admixed to cheaply available wheat bran. (6) The excellent effect is found for amylase production when 20% of wheat bran is admixed to defatted dry milk which is a poor medium. The activity is found to be $D^{40^{\circ}}_{30'}$ 7,000(L.S.V. 1,800) in 10% medium. (7) No significant effect is observed due to the addition of various inorganic salts. 3. Amylase production by solid cultures. Several conditions for amylase production by strain A-12 in wheat bran cultures have been studied and the results obtained are as follows. (1) The optimum conditions: are temperature $33^{\circ}C$, incubation lime 2 days, water content added 150 to 175% and the thickness of the medium 1.5cm, The activity is found to be $D^{40^{\circ}}_{30'}$ 36,000(L.S.V. 15,000) (2) No significant effect is found in the case of the additions of various organic and inorganic substances.

• Journal of Korea Foresty Energy
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• v.19 no.2
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• pp.93-101
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• 2000

Wood of Robinia pseudoacacia and bark of Populus alba$\times$P. glandulosa, Fraxinus rhynchophylla and Ulmus davidiana var. japonica were collected and extracted with acetone-water(7:3, v/v) in glass jar to examine whether its bioactive compounds exist. The concentrated extracts were fractionated with hexane, chloroform, ethylacetate and water, and then freeze-dried for column chromatography and bioactive tests. The isolated compounds were sakuranetin-5-O-$\beta$-D-glucopyranoside from Populus alba $\times$Pl glandulosa, 4--ethyoxy-(+)-leucorobinetinidin frm R. pseudoacacia and fraxetion from F. rhynchophylla and were characterized by $^1H$ and$^{13}C $ NMR and positive FAB-MS. Decay-resistant activity was expressed by weight loss ratio and hyphae growth inhibition in the wood dust agar medium inoculated wood rot fungi. R. pseudoacacia showed best anti-decaying property in both test and its methanol untreated samples, indicating higher activity than methanol treated samples in hyphae grwoth test. In antioxidative test, $\alpha$-tocopherol, one of natural antioxidants, and BHT, one of synthetic antioxidants, were used as references to cmpare with the antioxidant activities of the extacted fractions. Ethylacetate fraction of F. rhynchophylla bark indicated the hightest activity in this test and all fractions of R. pseudiacacia extractives also indicated higher activities compared with the other fractions. In the isolated compounds, aesculetin isolated from F. rhynchophylla bark showed best activity and followed by robonetinidin from R. pseudoacaica.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.28 no.6
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• pp.511-519
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• 2006

Autogenous bone grafts have been considered the gold standard for maxillofacial bony defects. However, this procedure could entail a complicated surgical procedure as well as potential donor site morbidity. Possibly the best solution for bone-defect regeneration is a tissue engineering approach, i.e. the use of a combination of a suitable scaffold with osteogenic cells. A major source of osteogenic cells is the bone marrow. Bone marrow-derived mesenchymal stem cells are multipotent and have the ability to differentiate into osteoblastic, chondrocytic, and adipocytic lineage cells. However, the isolation of cells from bone marrow has someproblems when used in clinical setting. Bone marrow aspiration is sometimes potentially more invasive and painful procedure and carries of a risk of morbidity and infection. A minimally invasive, easily accessible alternative would be cells derived from periosteum. The periosteum also contains multipotent cells that have the potential to differentiate into osteoblasts and chondrocytes. In the present study, we evaluated the osteogenic activity and mineralization of cultured human periosteal-derived cells. Periosteal explants were harvested from mandibule during surgical extraction of lower impacted third molar. The periosteal cells were cultured in the osteogenic inductive medium consisting of DMEM supplemented with 10% fetal calf serum, 50g/ml L-ascorbic acid 2-phosphate, 10 nmol dexamethasone and 10 mM -glycerophosphate for 42 days. Periosteal-derived cells showed positive alkaline phosphatase (ALP) staining during 42 days of culture period. The formation of ALP stain showed its maximal manifestation at day 14 of culture period, then decreased in intensity during the culture period. ALP mRNA expression increased up to day 14 with a decrease thereafter. Osteocalcin mRNA expression appeared at day 7 in culture, after that its expression continuously increased in a time-dependent manner up to the entire duration of culture. Von Kossa-positive mineralization nodules were first present at day 14 in culture followed by an increased number of positive nodules during the entire duration of the culture period. In conclusion, our study showed that cultured human periosteal-derived cells differentiated into active osteoblastic cells that were involved in synthesis of bone matrix and the subsequent mineralization of the matrix. As the periosteal-derived cells, easily harvested from intraoral procedure such as surgical extraction of impacted third molar, has the excellent potential of osteogenic capacity, tissue-engineered bone using periosteal-derived cells could be the best choice in reconstruction of maxillofacial bony defects.

• Microbiology and Biotechnology Letters
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• v.33 no.1
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• pp.16-23
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• 2005

In order to develop a new microbial fungicide using endophytic bacteria for the control of anthracnoses occurring on various crops, a total of 260 bacterial strains were isolated from fresh tissues of 5 plant species. After they were cultured in broth medium, their antifungal activities were tested for in vivo antifungal activity against cucumber anthracnose caused by Colletotrichum orbiculare. As the results, liquid cultures of 28 strains showed potent antifungal activities more than $90\%$ against cucumber anthracnose. At 3-fold dilutions of liquid cultures, 18 strains inhibited the development of cucumber anthracnose of more than $70\%$. They were further tested for in vivo antifungal activity against red pepper anthracnose caused by C. coccodes and in vitro antifungal activity against C. acutatum, a fungal agent causing red pepper anthracnose. Among 18 strains, a bacterial strain EB215 isolated from cucumber roots displayed the most potent antifungal activity against Colletotrichum species. It was identified as Burkholderia cepacia based on its physiological and biochemical characteristics, Biolog test and 16S rDNA gene sequence. It also controlled effectively the development of rice blast (Magnaporthe grisea), rice sheath blight (Corticium sasaki), tomato gray mold (Botrytis cinerea), and tomato late blight (Phytophthora infestans). Studies on the characterization of antifungal substances produced by B. cepacia EB215 are in progress.

• Microbiology and Biotechnology Letters
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• v.39 no.4
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• pp.317-323
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• 2011

Strain A-3, an amylase-producing bacteria, was isolated from coastal seawater near Daecheon in the Republic of Korea. It was seen to possess a single polar flagella and grow well, on ASW-YP agar plates, at temperatures of between $20-37^{\circ}C$. However, it grew more slowly at the temperatures of $15^{\circ}C$ and $40^{\circ}C$. Similarly, it was observed to grow abundantly, in an Artificial Sea Water-Yeast extract-Peptone (ASW-YP) liquid medium, in a pH range of 6-9, but not grow at pHs of 4-5 and a pH of 10. Strain A-3 was noted as being close to Pseudoalteromonas phenolica O-$BC30^T$, Pseudoalteromonas luteoviolacea $NCIMB1893^T$, Pseudoalteromonas rubra $ATCC29570^T$, and Pseudoalteromonas byunsanensis $FR1199^T$, with 98.30%, 97.86%, 97.78%, and 97.25% similarities respectively, in its 16S rRNA sequence. A phylogenetic tree revealed that strain A-3 and P. phenolica O-$BC30^T$ belong to a clade. However, strain A-3 differed from P. phenolica O-$BC30^T$ in relation to a number of physiological characteristics. Strain A-3 exhibited no growth above 5% NaCl concentrations, no utilization of D-glucose, D-mannose, D-maltose, or D-melibose, and no lipase (C-14) activity. All of these properties strongly indicate that strain A-3 is distant from P. phenolica O-$BC30^T$ and thus led us to name it Pseudoalteromonas sp. A-3. Pseudoalteromonas sp. A-3 produces ${\alpha}$-amylase throughout growth. Maximal amylase activities of 144.48 U/mL and 149.20 U/mL were seen at pH 7.0 and $37^{\circ}C$, respectively. Pseudoalteromonas sp. A-3's high, stable production of ${\alpha}$-amylase in addition to its biochemical features, such as alkalitolerance, suggest that it is a good candidate for industrial applications.

• Microbiology and Biotechnology Letters
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• v.41 no.2
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• pp.190-197
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• 2013

Three slime-forming lactic acid bacteria were isolated from traditional Korean fermented soy sauce and soybean paste and shown to produce exopolysaccharides (EPS) in sucrose media. By isolating the strains, examining their morphological characteristics and determining their 16S rDNA sequences, N58-5 and K6-7 were identified as Leuconostoc mesenteroides and N45- 10 as Leuconostoc citreum. The acid and bile tolerances of these three strains were investigated. Amongst the three lactic acid bacteria, Leuc. citreum N45-10 exhibited the highest viability ($10^5-10^6$ CFU/ml) in 0.05 M sodium phosphate buffer (pH 0.3) for 2 h, in artificial gastric juice for 2 h and in 0.3%, 0.5% oxgall for 24h. Leuc. mesenteroides K6-7, N58-5 and Leuc. citreum N45- 10 were grown in sucrose liquid medium and 8.16 g/L, 3.65 g/L, 16.17 g/L of EPS was collected, respectively. The hydrolyzed EPS was analyzed by HPLC in order to determine the sugar composition of EPS. Leuc. mesenteroides K6-7 and N58-5 showed two peaks indicating glucose and fructose, thus they were determined to be hetero-type polysaccharides. Leuc. citreum N45-10 showed only the glucose polymer, indicating it to be a homo-type polysaccharide. In addition, all three lactic acid bacterial hemolysis did not demonstrate a clear zone in blood agar in the area surrounding a lactic acid bacteria colony.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.32 no.4
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• pp.299-305
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• 2010

Purpose: Adipose tissue is located beneath the skin, around internal organs, and in the bone marrow in humans. Its main role is to store energy in the form of fat, although it also cushions and insulates the body. Adipose tissue also has the ability to dynamically expand and shrink throughout the life of an adult. Recently, it has been shown that adipose tissue contains a population of adult multipotent mesenchymal stem cells and endothelial progenitor cells that, in cell culture conditions, have extensive proliferative capacity and are able to differentiate into several lineages, including, osteogenic, chondrogenic, endothelial cells, and myogenic lineages. Materials and Methods: This study focused on endothelial cell culture from the adipose tissue. Adipose tissues were harvested from buccal fat pad during bilateral sagittal split ramus osteotomy for surgical correction of mandibular prognathism. The tissues were treated with 0.075% type I collagenase. The samples were neutralized with DMEM/and centrifuged for 10 min at 2,400 rpm. The pellet was treated with 3 volume of RBC lysis buffer and filtered through a 100 ${\mu}m$ nylon cell strainer. The filtered cells were centrifuged for 10 min at 2,400 rpm. The cells were further cultured in the endothelial cell culture medium (EGM-2, Cambrex, Walkersville, Md., USA) supplemented with 10% fetal bovine serum, human EGF, human VEGF, human insulin-like growth factor-1, human FGF-$\beta$, heparin, ascorbic acid and hydrocortisone at a density of $1{\times}10^5$ cells/well in a 24-well plate. Low positivity of endothelial cell markers, such as CD31 and CD146, was observed during early passage of cells. Results: Increase of CD146 positivity was observed in passage 5 to 7 adipose tissue-derived cells. However, CD44, representative mesenchymal stem cell marker, was also strongly expressed. CD146 sorted adipose tissue-derived cells was cultured using immuno-magnetic beads. Magnetic labeling with 100 ${\mu}l$ microbeads per 108 cells was performed for 30 minutes at $4^{\circ}C$ a using CD146 direct cell isolation kit. Magnetic separation was carried out and a separator under a biological hood. Aliquous of CD146+ sorted cells were evaluated for purity by flow cytometry. Sorted cells were 96.04% positivity for CD146. And then tube formation was examined. These CD146 sorted adipose tissue-derived cells formed tube-like structures on Matrigel. Conclusion: These results suggest that adipose tissue-derived cells are endothelial cells. With the fabrication of the vascularized scaffold construct, novel approaches could be developed to enhance the engineered scaffold by the addition of adipose tissue-derived endothelial cells and periosteal-derived osteoblastic cells to promote bone growth.

• Korean Journal of Environmental Biology
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• v.34 no.4
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• pp.304-313
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• 2016

One of the issues currently facing nuclear power plants is how to store spent nuclear waste materials which are contaminated with radionuclides such as $^{134}Cs$, $^{135}Cs$, and $^{137}Cs$. Bioremediation processes may offer a potent method of cleaning up radioactive cesium. However, there have only been limited reports on $Cs^+$ tolerant bacteria. In this study, we report the isolation and identification of $Cs^+$ tolerant bacteria in environmental soil and sediment. The resistant $Cs^+$ isolates were screened from enrichment cultures in R2A medium supplemented with 100 mM CsCl for 72 h, followed by microbial community analysis based on sequencing analysis from 16S rRNA gene clone libraries(NCBI's BlastN). The dominant Bacillus anthracis Roh-1 and B. cereus Roh-2 were successfully isolated from the cesium enrichment culture. Importantly, B. cereus Roh-2 is resistant to 30% more $Cs^+$ than is B. anthracis Roh-1 when treated with 50 mM CsCl. Growth experiments clearly demonstrated that the isolate had a higher tolerance to $Cs^+$. In addition, we investigated the adsorption of $0.2mg\;L^{-1}$ $Cs^+$ using B. anthracis Roh-1. The maximum $Cs^+$ biosorption capacity of B. anthracis Roh-1 was $2.01mg\;g^{-1}$ at pH 10. Thus, we show that $Cs^+$ tolerant bacterial isolates could be used for bioremediation of contaminated environments.

• Korean Journal of Environmental Agriculture
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• v.15 no.3
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• pp.362-371
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• 1996

This study was conducted to investigate the biochemical properties of isolated bacteria, low temperature tolerant methanogens which were selected for use as inoculum for anaerobic fermentation of agricultural and livestock wasted at low temperature. The results, obtained were summarized as follows: Low temperature tolerant methanogens were isolated from the samples which showed the high methanogenesis rate by enrichment culture at low temperature in methanol medium. These methanogens, Methanobacterium M-251 and Methanobacterium M-253 were isolated from swampy sediment at latitude $56.9^{\circ}$, Methanosarcina mazei M-372 from lake sediment IV at latitude $55.0^{\circ}$ N, and Methanobacterium formicicum M-375 from tidal land soil at latitude $37.0^{\circ}N$, respectively. The isolated anaerobic bacteria could not use sugars as carbon sources. The optimum pH value for the growth of M-251 and M-375 was 6.8, but those for M-253 and M-372 6.5 and 7.0, respectively. The minimum growth temperature of isolated, M-251 and M-253 were $8^{\circ}C$ and the optimum temperature $30^{\circ}C$, while the minimum of M-392 and M-395 were $13^{\circ}C$ and the optimum $37^{\circ}C$. The growth rate of isolates at $17.5^{\circ}C$ were lower by 32-50% than that of $30^{\circ}C$. The isolated Methanobacterium strains such as M-251, M-253, and M-375 have lower cell yield, 0.38-1.21g/1M $CH_4$ than 1.14-1.51g/1M $CH_4$ of Methanosarcina mazei M-372.