• 한국균학회소식:학술대회논문집
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• 2018.05a
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• pp.27-27
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• 2018

The production scale of mushroom cultivation in Korea is approximately 600 billion won, which is 1.6% of the Korean gross agricultural output. Annually, ca. 190,000 tons of mushrooms are harvested in Korea. Although the numbers of mushroom farms and cultivators are constantly decreasing, the total mushroom yields are increasing due to the large-scale cultivation facilities and automation. The recent expansion of the well-being trend causes increase in mushroom consumption in Korea: annual per capita consumption of mushroom was 3.9kg ('13) that is a little higher than European's average. Thus the exports of mushrooms, mainly Flammulina velutipes and Pleurotus ostreatus, have been increased since the middle of 2000s. Recently, however, it is slightly reduced. However, Vietnam, Hong Kong, the United States, the Netherlands and continued to export, and the country has increased recently been exported to Australia, Canada, Southeast Asia and so on. Canned foods of Agaricus bisporus was the first exports of the Korean mushroom industry. This business has reached the peak of the sale in 1977-1978. As Korea initiated trade with China in 1980, the international prices of mushrooms were sharply fall that led to shrink the domestic markets. According to the high demand to develop new items to substitute for A. bisporus, oyster mushroom (Pleurotus ostreatus) was received the attention since it seems to suit the taste of Korean consumers. Although log cultivation technique was developed in the early 1970s for oyster mushroom, this method requires a great deal of labor. Thus we developed shelf cultivation technique which is easier to manage and allows the mass production. In this technique, the growing shelf is manly made from fermented rice straw, that is the unique P. ostreatus medium in the world, was used only in South Korea. After then, the use of cotton wastes as an additional material of medium, the productivity. Currently it is developing a standard cultivation techniques and environmental control system that can stably produce mushrooms throughout the year. The increase of oyster mushroom production may activate the domestic market and contribute to the industrial development. In addition, oyster mushroom production technology has a role in forming the basis of the development of bottle cultivation. Developed mushroom cultivation technology using bottles made possible the mass production. In particular, bottle cultivation method using a liquid spawn can be an opportunity to export the F.velutipes and P.eryngii. In addition, the white varieties of F.velutipes were second developed in the world after Japan. We also developed the new A.bisporus cultivar "Sae-ah" that is easy to grown in Korea. To lead the mushroom industry, we will continue to develop the cultivars with an international competitive power and to improve the cultivation techniques. Mushroom research in Korea nowadays focuses on analysis of mushroom genetics in combination with development of new mushroom varieties, mushroom physiology and cultivation. Further studied are environmental factors for cultivation, disease control, development and utilization of mushroom substrate resources, post-harvest management and improvement of marketable traits. Finally, the RDA manages the collection, classification, identification and preservation of mushroom resources. To keep up with the increasing application of biotechnology in agricultural research the genome project of various mushrooms and the draft of the genetic map has just been completed. A broad range of future studies based on this project is anticipated. The mushroom industry in Korea continually grows and its productivity rapidly increases through the development of new mushrooms cultivars and automated plastic bottle cultivation. Consumption of medicinal mushrooms like Ganoderma lucidum and Phellinus linteus is also increasing strongly. Recently, business of edible and medicinal mushrooms was suffering under over-production and problems in distribution. Fortunately, expansion of the mushroom export helped ease the negative effects for the mushroom industry.

• Korean Journal of Microbiology
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• v.52 no.1
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• pp.10-17
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• 2016

Enzyme immunoassay to analyze specific binding activity of antibody to antigen uses horseradish peroxidase (HRP) or alkaline phosphatase (AP). Chemical methods are usually used for coupling of these enzymes to antibody, which is complicated and random cross-linking process. As results, it causes decreases or loss of functional activity of either antibody or enzyme. In addition, most enzyme assays use secondary antibody to detect antigen binding activity of primary antibody. Enzymes coupled to secondary antibody provide a binding signal by substrate-based color development, suggesting secondary antibody is required in enzyme immunoassay. Additional incubation time for binding of secondary antibody should also be necessary. More importantly, non-specific binding activity caused by secondary antibody should also be eliminated. In this study, we cloned AP isolated from Escherichia coli (E. coli) chromosome by PCR and fused to) hAY4 single-chain variable domain fragment (ScFv) specific to death receptor (DR4) which is a receptor for tumor necrosis factor ${\alpha}$ related apoptosis induced ligand (TRAIL). hAY4 ScFv-AP expressed in E. coli showed 73.8 kDa as a monomer in SDS-PAGE. However, this fusion protein shown in size-exclusion chromatography (SEC) exhibited 147.6 kDa as a dimer confirming that natural dimerization of AP by non-covalent association induced ScFv-AP dimerization. In several immunoassay such as ELISA, Western blot and immunocytochemistry, it showed antigen binding activity by color development of substrates catalyzed by AP directly fused to primary hAY4 ScFv without secondary antibody. In summary, hAY4 ScFv-AP fusion protein was successfully purified as a soluble dimeric form in E. coli and showed antigen binding activity in several immunoassays without addition of secondary antibody which sometimes causes time-consuming, expensive and non-specific false binding.

• Korean Journal of Environment and Ecology
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• v.33 no.5
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• pp.525-538
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• 2019

The purpose of this study was to analyze the habitat status and species diversity of benthic diatoms and estimate the applicability of TDI (Trophic Diatom Index) to obtain the basic data for the identification and management of created wetlands in the Nakdong River. We observed a total of 38 families and 173 species of benthic diatom during the survey period, and spring and autumn showed a similar number of species of 156 and 154, respectively. The result of the SOM (Self-Organizing Map) analysis showed that the distribution of benthic diatom was sensitive to environmental factors such as nutrient concentration and rainfall in each wetland. The cluster 1 was characterized by the survey sites of autumn mostly and consisted of points of high TDI, although the nutrients such as total phosphorus and total nitrogen were low, and the species number and abundance of diatoms were low. Conversely, cluster 4 was characterized by the survey sites of spring mostly and consisted of points of low TDI, even though total nitrogen was high. Considering that most of the created wetlands had the reduced inflow and outflow, the increased flow rate in the summer lowers nutrient values in autumn, and the species number and abundance of benthic diatom decreases due to the increase of turbidity, which reduces the light penetrations to the substrates. On the contrary, the TDI value is low in spring because the low water level causes insufficient substrate surface to the benthic diatoms, and it is too early for the establishment and development of saprophilous species. Although various studies have used TDI as an indicator for evaluating the habitat environment and water quality, it is not a good evaluation indicator in this study since the nutrient concentration in the wetlands mostly high as they have a low flow rate and are close to the stagnant area. Nevertheless, additional periodic surveys that comprehensively reflect the fact that the summer rainfall and inflow/outflow regulating function might affect the species diversity and distribution of benthic diatoms are necessary.

• 한국균학회소식:학술대회논문집
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• 2016.05a
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• pp.19-20
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• 2016

Sesquiterpenoids are defined as $C_{15}$ compounds derived from farnesyl pyrophosphate (FPP), and their complex structures are found in the tissue of many diverse plants (Degenhardt et al. 2009). FPP's long chain length and additional double bond enables its conversion to a huge range of mono-, di-, and tri-cyclic structures. A number of cyclic sesquiterpenes with alcohol, aldehyde, and ketone derivatives have key biological and medicinal properties (Fraga 1999). Fungi, such as the wood-rotting Polyporus brumalis, are excellent sources of pharmaceutically interesting natural products such as sesquiterpenoids. In this study, we investigated the biosynthesis of P. brumalis sesquiterpenoids on modified medium. Fungal suspensions of 11 white rot species were inoculated in modified medium containing $C_6H_{12}O_6$, $C_4H_{12}N_2O_6$, $KH_2PO_4$, $MgSO_4$, and $CaCl_2$ for 20 days. Cultivation was stopped by solvent extraction via separation of the mycelium. The metabolites were identified as follows: propionic acid (1), mevalonic acid lactone (2), ${\beta}$-eudesmane (3), and ${\beta}$-eudesmol (4), respectively (Figure 1). The main peaks of ${\beta}$-eudesmane and ${\beta}$-eudesmol, which were indicative of sesquiterpene structures, were consistently detected for 5, 7, 12, and 15 days These results demonstrated the existence of terpene metabolism in the mycelium of P. brumalis. Polyporus spp. are known to generate flavor components such as methyl 2,4-dihydroxy-3,6-dimethyl benzoate; 2-hydroxy-4-methoxy-6-methyl benzoic acid; 3-hydroxy-5-methyl phenol; and 3-methoxy-2,5-dimethyl phenol in submerged cultures (Hoffmann and Esser 1978). Drimanes of sesquiterpenes were reported as metabolites from P. arcularius and shown to exhibit antimicrobial activity against Gram-positive bacteria such as Staphylococcus aureus (Fleck et al. 1996). The main metabolites of P. brumalis, ${\beta}$-Eudesmol and ${\beta}$-eudesmane, were categorized as eudesmane-type sesquiterpene structures. The eudesmane skeleton could be biosynthesized from FPP-derived IPP, and approximately 1,000 structures have been identified in plants as essential oils. The biosynthesis of eudesmol from P. brumalis may thus be an important tool for the production of useful natural compounds as presumed from its identified potent bioactivity in plants. Essential oils comprising eudesmane-type sesquiterpenoids have been previously and extensively researched (Wu et al. 2006). ${\beta}$-Eudesmol is a well-known and important eudesmane alcohol with an anticholinergic effect in the vascular endothelium (Tsuneki et al. 2005). Additionally, recent studies demonstrated that ${\beta}$-eudesmol acts as a channel blocker for nicotinic acetylcholine receptors at the neuromuscular junction, and it can inhibit angiogenesis in vitro and in vivo by blocking the mitogen-activated protein kinase (MAPK) signaling pathway (Seo et al. 2011). Variation of nutrients was conducted to determine an optimum condition for the biosynthesis of sesquiterpenes by P. brumalis. Genes encoding terpene synthases, which are crucial to the terpene synthesis pathway, generally respond to environmental factors such as pH, temperature, and available nutrients (Hoffmeister and Keller 2007, Yu and Keller 2005). Calvo et al. described the effect of major nutrients, carbon and nitrogen, on the synthesis of secondary metabolites (Calvo et al. 2002). P. brumalis did not prefer to synthesize sesquiterpenes under all growth conditions. Results of differences in metabolites observed in P. brumalis grown in PDB and modified medium highlighted the potential effect inorganic sources such as $C_4H_{12}N_2O_6$, $KH_2PO_4$, $MgSO_4$, and $CaCl_2$ on sesquiterpene synthesis. ${\beta}$-eudesmol was apparent during cultivation except for when P. brumalis was grown on $MgSO_4$-free medium. These results demonstrated that $MgSO_4$ can specifically control the biosynthesis of ${\beta}$-eudesmol. Magnesium has been reported as a cofactor that binds to sesquiterpene synthase (Agger et al. 2008). Specifically, the $Mg^{2+}$ ions bind to two conserved metal-binding motifs. These metal ions complex to the substrate pyrophosphate, thereby promoting the ionization of the leaving groups of FPP and resulting in the generation of a highly reactive allylic cation. Effect of magnesium source on the sesquiterpene biosynthesis was also identified via analysis of the concentration of total carbohydrates. Our current study offered further insight that fungal sesquiterpene biosynthesis can be controlled by nutrients. To profile the metabolites of P. brumalis, the cultures were extracted based on the growth curve. Despite metabolites produced during mycelia growth, there was difficulty in detecting significant changes in metabolite production, especially those at low concentrations. These compounds may be of interest in understanding their synthetic mechanisms in P. brumalis. The synthesis of terpene compounds began during the growth phase at day 9. Sesquiterpene synthesis occurred after growth was complete. At day 9, drimenol, farnesol, and mevalonic lactone (or mevalonic acid lactone) were identified. Mevalonic acid lactone is the precursor of the mevalonic pathway, and particularly, it is a precursor for a number of biologically important lipids, including cholesterol hormones (Buckley et al. 2002). Farnesol is the precursor of sesquiterpenoids. Drimenol compounds, bi-cyclic-sesquiterpene alcohols, can be synthesized from trans-trans farnesol via cyclization and rearrangement (Polovinka et al. 1994). They have also been identified in the basidiomycota Lentinus lepideus as secondary metabolites. After 12 days in the growth phase, ${\beta}$-elemene caryophyllene, ${\delta}$-cadiene, and eudesmane were detected with ${\beta}$-eudesmol. The data showed the synthesis of sesquiterpene hydrocarbons with bi-cyclic structures. These compounds can be synthesized from FPP by cyclization. Cyclic terpenoids are synthesized through the formation of a carbon skeleton from linear precursors by terpene cyclase, which is followed by chemical modification by oxidation, reduction, methylation, etc. Sesquiterpene cyclase is a key branch-point enzyme that catalyzes the complex intermolecular cyclization of the linear prenyl diphosphate into cyclic hydrocarbons (Toyomasu et al. 2007). After 20 days in stationary phase, the oxygenated structures eudesmol, elemol, and caryophyllene oxide were detected. Thus, after growth, sesquiterpenes were identified. Per these results, we showed that terpene metabolism in wood-rotting fungi occurs in the stationary phase. We also showed that such metabolism can be controlled by magnesium supplementation in the growth medium. In conclusion, we identified P. brumalis as a wood-rotting fungus that can produce sesquiterpenes. To mechanistically understand eudesmane-type sesquiterpene biosynthesis in P. brumalis, further research into the genes regulating the dynamics of such biosynthesis is warranted.