• 한국균학회소식:학술대회논문집
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• 2015.11a
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• pp.37-37
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• 2015

dsRNA was found in malformed cultures of Lentinula edodes strain FMRI0339, one of the three most popular sawdust cultivated commercial strains of shiitake, and was also found in healthy-looking fruiting bodies and actively growing mycelia. Cloning of the partial genome of the dsRNA revealed the presence of the RdRp sequence of a novel L. edodes mycovirus (LeV), and sequence comparison of the cloned amplicon showed an identical sequence to known RdRp genes of LeV found in strain HKA. The meiotic stability of dsRNA was examined by measuring the ratio of the presence of dsRNA among sexual monokaryotic progeny. More than 40% of the monokaryotic progeny still contained the dsRNA, indicating the persistence of dsRNA during sexual reproduction. Comparing the mycelia growth of monokaryotic progeny suggested that, although variations in the growth rate existed among progeny and virus infection was observed in highly actively growing progeny, there appeared to be a tendency toward a lower frequency of virus incidence in actively growing progeny. This study attempted to cure the edible mushroom L. edodes strain FMRI0339 of the L. edodes mycovirus (LeV) in order to obtain an isogenic virus-free fungal strain as well as a virus-infected strain for comparison. Mycelial fragmentation, followed by being spread on a plate with serial dilutions resulted in a virus-free colony. Viral absence was confirmed with gel electrophoresis after dsRNA-specific virus purification, Northern blot analysis, and PCR using reverse transcriptase (RT-PCR). Once cured, all of fungal cultures remained virus-free over the next two years. Interestingly, the viral titer of LeV varied depending on the culture condition. The titer from the plate culture showed at least a 20-fold higher concentration than that grown in the liquid culture. However, the reduced virus titer in the liquid culture was recovered by transferring the mycelia to a plate containing the same medium. In addition, oxygen-depleted culture conditions resulted in a significant decrease of viral concentration, but not to the extent seen in the submerged liquid culture. Although no $discernable phenotypic changes in colony morphology were observed, virus-cured strains showed significantly higher growth rates and mycelial mass than virus-infected strains. We were also explored effects of LeV on fruiting body formation and mushroom yield. The fruiting body formation yield of virus-free L. edodes was larger than virus-infected L. edodes. These results indicate that LeV infection has a deleterious effect on mycelial growth and fruiting body formation. In addition, we have been investigated host-parasite interaction between L. edodes and its mycovirus interaction to study viral mechanism by establishment of proteomics.

• Journal of the East Asian Society of Dietary Life
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• v.15 no.4
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• pp.386-396
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• 2005

The effects of spirulina-added salad dressing on lipid profiles and antioxidant biomarkers such as total glutathionine, TBARS value, carbonyl value, GPx, GR, SOD and paraoxonase activity in plasma or liver of mice were evaluated Sixteen male ICR mice weighing 20$\pm$2 g were divided into two groups and fed low fat ($5\%$ fat) diet (low fat control: LFC) and low fat control plus dressing diet (LFD) for eight weeks. Body weight, tissue weights of liver, heart and kidney, and the distribution of body fat deposition were not significantly different between two groups. Also, the profile of TG, TC, LDL and HDL cholesterol were similar between two groups. The DNA damage was determined using the comet assay (single cell gel assay) with alkaline electrophoresis and quantified by measuring tail length (TL). Spirulina salad dressing consumption resulted in significant decrease in lymphocyte DNA damage expressed by TL (LFC: $28.8{\mu}m$, LFD: $20.3{\mu}m$). Additionally, salad dressing consumption for 8 wks decreased the lipid peroxidation assayed by TBARS to $12.6\%$ compared with the control. The levels of antioxidant vitamins such as $\beta$-carotene were significantly higher in plasma of LFD group than those in LFC group based on HPLC method This study shows that spirulina-added salad dressing exerts degenerative disease-protective effects on oxidative DNA damage and lipid peroxidation possibly via a free radical levels.

• The Korean Journal of Zoology
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• v.33 no.1
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• pp.63-69
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• 1990

To study the follicular atretic mechanism in mammalian ovary, the medium-sized (3.0-6.0mm) follicles of porcine ovary were morphologically classified as nonnal and atretic. Steroid concentrations in the follicular fluid were analyzed by radioimmunoassay, and the proteins in that fluid were electrophoretically separated. Concentrations of progesterone in the atretic follicular fluid of follicular phases were higher than those of normal ones (p < 0.05). Concentrations of testosterone were high in normal luteal and atretic follicular-phases follicles. The concentrations of estradiol remained significanily lower in atretic follicular-phases follicles than normal. After sodium dodecyl sulfate-polyacrylamide gel electrophoresis of follicular fluid proteins, four kinds of proteins (20K, 32K, 33K, 38K) were detected, and those proteins were not present in sera. According to the ovarian cycle, proteins of MW of 112K and 141K were identified. In atretic follicular fludies, MW of 23K and 24K were specifically detected. From the above results, we can conclude that, as ovarian cycle changes, steroid contents and protein composition in atretic follicular fluid are different from the normal developing follicular fluid. To further understand the physiologic roles of the proteins present in the atretic follicular fluids, these proteins need to be characterized and identified.

• Journal of Veterinary Clinics
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• v.25 no.5
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• pp.340-345
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• 2008

Malassezia pachydermatis (M. pachydermatis) is a component of the normal cutaneous flora of the dog and atopic dermatitis (AD) is one of the most common diseases associated with Malassezia overgrowth in dogs. The purpose of this study was to investigate the humoral response (IgG) to extracts of M. pachydermatis of in a dog with AD. We used Western immunoblotting to identify allergens of M. pachydermatis. Gel electrophoresis of extracts proteins and immunoblotting of sera samples in both an atopic dog and a non-atopic dog were compared. Proteins of 18, 21, 26, 32, 34, 38, 40, 42, 46, 58, 64, 75, 85, and 120 kDa were observed in a serum of atopic dog. However, when serum of a non-atopic dog was used, protein bands were not identified except binding in 50 kDa protein. The results of this study indicate that atopic dogs with M. pachydermatis dermatitis may induce IgG response and also suggest that humoral response to M. pachydermatis could be important in pathogenesis of AD in dogs. However, further studies are required to identify roles of humoral response to M. pachydermatis in canine AD.

• Journal of Veterinary Clinics
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• v.27 no.5
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• pp.508-513
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• 2010

Cytokines are important mediators of the immune response, and quantitating cytokine mRNA is a highly sensitive and attractive method for measuring cytokine production. The objective of the current study was to develop and validate a SYBR green quantitative real-time reverse transcriptase PCR (qRT-PCR) assay for measuring canine cytokine mRNA. The optimal annealing temperatures ($T_a$) of the designed primers were $62^{\circ}C$ for interleukin (IL)-$1{\beta}$, IL-6 and IL-10; $60^{\circ}C$ for glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and tumor necrosis factor (TNF)-${\alpha}$; and $58^{\circ}C$ for high mobility group box 1 (HMGB1). Primer efficiencies of all primers calculated for standard curve samples were between 97.1% and 102.6%. No evidence of secondary structure or primer-dimer formation was seen via melt-curve analysis or gel electrophoresis. The developed qRT-PCR assays are highly specific and sensitive and can be used to quantify gene expression levels of canine cytokines.

• Journal of Life Science
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• v.25 no.6
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• pp.673-679
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• 2015

The culture conditions needed to maximize the production of laccase from Pholiota highlandensis mycelia were investigated. Among the tested media for laccase production, Coriolus versicolor medium (CVM; 2% dextrose, 0.4% peptone, 0.6% yeast extract, 0.046% KH₂PO₄, 0.1% K₂HPO₄, 0.05% MgSO₄·7H₂O) showed the highest activity for the enzyme. Then, to optimize culture conditions for laccase activity, the influences of various carbon, nitrogen, phosphorus, and inorganic salt sources in CVM were investigated. The optimum culture medium was 2% fructose, 0.4% peptone with 0.6% yeast extract, 0.05% NaH₂PO₄, and 0.05% MgSO₄·7H₂O as carbon, nitrogen, phosphorus, and inorganic salt sources, respectively. Several aromatic compounds in the medium enhanced laccase activity to varying degrees. Guaiacol induced maximum laccase production, yielding 114.1 U/ml laccase activity after cultivation for 11 days at 25℃. The optimum pH and temperature for laccase production were 8.0 and 35℃, respectively. Native polyacrylamide-gel electrophoresis (PAGE) followed by laccase-activity staining with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) as the substrate was performed to identify the presence of laccase under the optimum conditions studied. Zymogram analysis of the supernatant culture showed an enzymatic band with a molecular mass of about 90 kDa.

• Journal of Life Science
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• v.25 no.2
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• pp.197-205
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• 2015

The comparative effects of the fibrinolytic action, antioxidative activity, and tyrosinase inhibition of Cordyceps militaris powder and fermented Cordyceps militaris powders were investigated using several microscopic organisms. The nutritional components such as phenolic compounds, flavonoids, and minerals were also measured. The total phenolic compounds and flavonoid concentrations were highest in the Cordyceps militaris powder fermented by Aspergillus oryzae. Major minerals were K, Ca, Mg, and Zn. Native polyacrylamide gel electrophoresis (native-PAGE) analysis of the total protein patterns of Cordyceps militaris powder and fermented Cordyceps militaris powders revealed slight varietal differences. Fibrinolytic activity was highest in the Cordyceps militaris powder fermented by Bacillus subtilis and Aspergillus kawachii. The DPPH radical scavenging activity was slightly stronger in the powder fermented by Monascus purpureus; however, these samples all exhibited a relatively low activity when compared with butylated hydroxytoluene (BHT). Tyrosinase inhibition activity was stronger in the powder fermented by Aspergillus oryzae than in unfermented powder. These results may provide basic data for understanding the biological activities and chemical characteristics of Cordyceps militaris powder fermented by several microscopic organisms for the development of functional foods.

• Journal of Life Science
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• v.22 no.11
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• pp.1523-1528
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• 2012

In a previous proteomic study, heat shock protein beta 1 (HSPB1) was detected as differentially expressed protein in longissimus thoracis between low (grade 3) and high (grade 1++) meat quality groups by 2DE gel electrophoresis. The present study investigated an association of HSPB1 expression at the level of gene and protein with Warner-Bratzler shear force (WBS) measured in 20 Hanwoo steers. An analysis of variance (ANOVA) between expression values and WBS showed that WBS was affected by HSPB1 expression (p<0.05). The expression (at both gene and protein level) of the HSPB1 was 2 times higher in the low WBS group than that in the high WBS group (p<0.01). This result suggests that the HSPB1 gene may be a candidate gene associated with tenderness in longissimus thoracis of Korean cattle.

• Journal of Life Science
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• v.18 no.3
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• pp.336-343
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• 2008

Histone deacetylases (HDACs) inhibitors have emerged as the accessory therapeutic agents for various human cancers, since they can block the activity of specific HDACs, restore the expression of some tumor suppressor genes and induce cell differentiation, cell cycle arrest and apoptosis in vitro and in vivo. In the present study, we investigated that the effect of trichostatin A (TSA), an HDAC inhibitor, on the cell growth and apoptosis, and its effect on the nuclear factor-kappaB $(NF-{\kappa}B)$ activity in 267B1 human prostate epithelial cells. Exposure of 267B1 cells to TSA resulted in growth inhibition and apoptosis induction in and dose-dependent manners as measured by fluorescence microscopy, agarose gel electrophoresis and flow cytometry analysis. TSA treatment inhibited the levels of IAP family members such as c-IAP-1 and c-IAP-2 and induced the proteolytic activation of caspase-3, -8 and -9, which were associated with concomitant degradation of poly (ADP-ribose)-polymerase, ${\beta}-catenin$ and laminin B proteins. The increase in apoptosis by TSA was connected with the translocation of $NF-{\kappa}B$ from cytosol to nucleus, increase of the DNA binding as well as promoter activity of $NF-{\kappa}B$, and degradation of cytosolic inhibitor of KappaB $(I{\kappa}B)-{\alpha}$ protein. We therefore concluded that TSA demonstrated anti-proliferative and apoptosis-inducing effects on 267B1 cells in vitro, and that the activation of caspases and $NF-{\kappa}B$ may play important roles in its mechanism of action. Although further studies are needed, these findings provided important insights into the possible molecular mechanisms of the anti-cancer activity of TSA.

• Journal of Life Science
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• v.26 no.9
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• pp.1007-1014
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• 2016

A highly efficient, rapid, and reliable multiplex polymerase chain reaction based method for distinguishing ten species of genus Cynoglossus (C. senegalensis, C. abbreviates, C. macrolepidotus, C. arel, C. semilaevis, C. interruptus, C. joyneri, C. lingua, C. robustus, and C. monodi) is described. The species-specific primer sets were designed base on the cytochrome oxidase subunit I gene (1,500 bp). The optimal PCR conditions and primers were selected for ten of Cynoglossus species to determine target base sequences using single PCR. Multiplex PCR using the ten pairs of primers either specifically amplified a DNA fragment of a unique size or failed, depending on each species DNA. The length of amplification fragment of 208 bp for C. senegalensis, 322 bp for C. abbreviates, 493 bp for C. macrolepidotus, 754 bp for C. arel, 874 bp for C. semilaevis, 952 bp for C. interruptus, 1,084 bp for C. joyneri, 1,198 bp for C. lingua, 1,307 bp for C. robustus, and 1,483 bp for C. monodi with the species-specific primers, visualized by agarose gel electrophoresis, allowed perfectly distinction of the Cynoglossus species. The multiplex PCR assay can be easily performed on multiple samples and attain final results in less than 6 hours. This technique should be a useful addition to the molecular typing tools for the tentative identification of Cynoglossus species.