• Korean Chemical Engineering Research
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• 제47권2호
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• pp.220-229
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• 2009

본 연구에서는 Phellinus linteus 균사체의 액상배양을 통한 면역증강 생리활성 효능의 단백다당체 생산공정을 개발하기 위한 시도로서, 우선 생산균체의 원형질체 형성을 통한 고생산성 균주를 개발하고자 하였으며, 발효기 액상배양 시 최적 배양형태의 유도를 통해 균사체와 단백다당체의 생산성을 극대화하고자 하였다. 본 연구실에서 채취한 생산 균주를 ITS rDNA sequencing 방법과 blast search 방법에 의해 조사한 결과 다양한 Phellinus linteus 종들과 99.67% 이상의 유사성 확인되어, 이 균주를 Phellinus linteus라고 최종적으로 동정할 수 있었다. 이 동정된 균주로부터 균주 개량을 시도하기 위해 Phellinus linteus 균사체로부터 대량의 원형질체 형성 및 재생에 의한 단일 콜로니 획득 방법을 개발함으로써 균주를 신속하게 개량할 수 있었다. Sorbitol을 이용한 banding filtration 방법을 이용하여 원형질체를 회수한 결과 $10^5{\sim}10^6\;protoplasts/ml$를 얻을 수 있었으며, 원형질체 재생률은 $10^{-2}{\sim}10^{-3}$로 나타났다. 균주개량을 위해 원형질체 재생배지와 고체배양배지에서 고성장성 및 고안정성을 보이는 균주들을 지속적으로 대량 선별하여, 액상 생산배양을 수행하였다. 그 결과 균사체량은 13~15 g/L로 대부분 비슷하게 자랐으며, 조단백다당체의 함량 또한 5.8~6.4%로 거의 비슷하게 분포하는 것으로 나타났는데, 이로부터 고체배양배지에서 빠른 성장속도를 보여주는 균주들이 대부분 액상 생산배양에서도 고생산성 및 고안정성을 보여주는 것을 확인할 수 있었다. 한편 Phellinus linteus 균사체의 경우 조단백다당체의 함량이 세포 무게당 거의 일정한 양을 함유하고 있는 것으로 확인되었으므로, 조단백다당체의 생산성을 증가시키기 위해서는 최종 생산배양에서의 균체량 증가가 가장 중요한 것으로 판단되어, 균사형성 고등균류의 균사체 배양 시 균체량 증가에 가장 중요한 요인 중의 하나인 생산균주의 배양형태적 특성에 대해 집중적으로 조사하였다. 균주개량 실험을 통해 고생산성 균주로 최종 결정된 AR147 균주를 이용해서 다양한 배양조건에서 발효조 배양을 수행한 결과, 최종 생산발효조로의 접종원이 고농도의 균사모양인 경우에 생산균주의 배양형태가 매우 작은 compact한 펠렛 모양(대부분 직경 0.5 mm 이하)을 유지하는, 이상적인 균사체 액상배양 공정이 이루어지는 것으로 확인되었다. 즉 생산 발효조배양에서 직경 0.5 mm 이하의 compact한 펠렛 모양의 배양형태가 유도되었을 경우, lag phase 시간의 획기적 감소와 1.5배 이상의 높은 세포비성장속도로 인해, 최종 균사체생산성이 다른 배양형태를 유도한 경우에 비해 약 3.3배 더 높은 주목할 만한 배양결과를 얻을 수 있었다. 이로부터 균사 형성 Phellinus linteus의 산업용 발효조 배양 시, 각 배양단계에서의 생산균체의 배양형태가 최종 균체생산성, 궁극적으로는 최종 단백다당체의 생산성에 심각한 영향을 미친다는 것을 알 수 있었다.

• 한국초지조사료학회지
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• 제36권3호
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• pp.237-242
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• 2016

Copper (Cu) is a necessary microelement for plants. However, high concentrations of Cu are toxic to plants that change the regulation of several stress-induced proteins. In this study, an annealing control primer (ACP) based approach was used to identify differentially expressed Cu-induced genes in alfalfa leaves. Two-week-old alfalfa plants (Medicago sativa L.) were exposed to Cu for 6 h. Total RNAs were isolated from treated and control leaves followed by ACP-based PCR technique. Using GeneFishing ACPs, we obtained several genes those expression levels were induced by Cu. Finally, we identified several genes including UDP-glucuronic acid decarboxylase, transmembrane protein, small heat shock protein, C-type cytochrome biogenesis protein, mitochondrial 2-oxoglutarate, and trans-2,3-enoyl-CoA reductase in alfalfa leaves. These identified genes have putative functions in cellular processes such as cell wall structural rearrangements, transduction, stress tolerance, heme transport, carbon and nitrogen assimilation, and lipid biosynthesis. Response of Cu-induced genes and their identification in alfalfa would be useful for molecular breeding to improve alfalfa with tolerance to heavy metals.

• Asian-Australasian Journal of Animal Sciences
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• 제17권6호
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• pp.777-783
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• 2004

Samples of 13 species of forest tree leaves fed to livestock in the semi-hilly arid zone of Punjab State in India were collected at 30 d interval for 12 months, in order to assess their nutritional worth for livestock. The ground samples were pooled for 4 different seasons viz. dry hot, hot humid, fall and winter. The chemical composition irrespective of the season revealed that CP content varied between 8.9 (Carrisa) to 22.0% (Leucaena). Globulin was the major protein fraction in most of the leaves. The lowest concentration of cell wall constituents was observed in Morus alba and Grewea. The leaves in general became fiberous and lignified during winter and fall as compared to summer season. The leaves of Grewea, Morus alba, Leucaena, Carrisa and Acacia were rich in Ca, P and most of the trace elements. The total phenolics ranged between 1.88% (Azardirachta) to 15.82% (Acacia). The leaves of Acacia had the highest concentration of hydrolysable tannins (14.6%) whereas that of Carrisa had that of condensed tannins (5.9%). The condensed tannins (more than 3%) were negatively correlated to the digestibility of dry matter (DM), neutral detergent fiber (NDF) and crude protein (CP). The digestion kinetic parameters for DM, NDF and CP revealed that leaves of Morus alba, Zizyphus and Ehretia had highest insoluble but potentially degradable fraction. The minimum rumen fill values also revealed that leaves of Grewea, Azardirachta, Morus, Ehretia and Leucaena had great potential for voluntary DM intake. The leaves of Ougeinia, Malha, Dodenia and Carrisa had significantly higher rumen fill value indicating poor potential for voluntary DM intake. Season did not have any significant impact on digestion kinetic parameters except that most of the leaves had low potentially degradable fraction, which was degraded at slow rate during winter. It was concluded that the leaves of Morus, Ehretia, Grewea and Leucaena had great potential as livestock feed, while feeding of Ougeinia, Malha and Dodonea leaves should be avoided.

• Asian-Australasian Journal of Animal Sciences
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• 제29권1호
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• pp.51-61
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• 2016

In order to provide recommendations on the most useful forage species to smallholder farmers, eleven grass and eleven legume forages grown in Abomey-Calavi in Republic of Benin were investigated for nutritive value (i.e. chemical composition and energy content) and fermentation characteristics (i.e. gas and volatile fatty acid production, organic matter degradability). The in vitro gas production technique was used, incubating the forages for 120 h under anaerobic condition with buffalo rumen fluid. Compared to legume, tropical grass forages showed lower energy (8.07 vs 10.57 MJ/kg dry matter [DM]) and crude protein level (16.10% vs 19.91% DM) and higher cell wall content (neutral detergent fiber: 63.8% vs 40.45% DM), respectively. In grass forages, the chemical composition showed a quite high crude protein content; the in vitro degradability was slightly lower than the range of tropical pasture. The woody legumes were richer in protein and energy and lower in structural carbohydrates than herbaceous plants, however, their in vitro results are influenced by the presence of complex compounds (i.e. tannins). Significant correlations were found between chemical composition and in vitro fermentation characteristics. The in vitro gas production method appears to be a suitable technique for the evaluation of the nutritive value of forages in developing countries.

• Asian-Australasian Journal of Animal Sciences
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• 제9권2호
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• pp.133-137
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• 1996

Duckweed, an aquatic plant of the family Lemnaceae, is a rich source of protein and also contains cell wall materials. Spirodela, Lemna and Wolffia, the most available species of duckweeds were evaluated in terms of their chemical composition, the rate and extent of digestion of their dry matter(DM) and crude protein(CP) in the rumen and also their acceptability to the cattle. The three species contained CP of 284, 399 and $299g{\cdot}kg^{-1}$ DM, respectively; NDF of 471, 574 and $476g{\cdot}kg^{-1}$ DM, respectively; ADF of 215, 203 and $227g{\cdot}kg^{-1}$ DM, respectively. The rumen digestibilities of DM of the three species for 24 h were 410, 570 and $731g{\cdot}kg^{-1}$ DM, respectively and of CP were 528, 740 and $778g{\cdot}kg^{-1}$ DM, respectively. The rates of digestion of DM of the three duckweeds were 2.22, 3.63 and $5.73%h^{-1}$, respectively and of CP were 5.14, 4.22 and $6.05%h^{-1}$, respectively. Similarly, the extent of digestion of DM were 853, 723 and $926g{\cdot}kg^{-1}$ DM, respectively and of CP were 801, 874 and $943g{\cdot}kg^{-1}$ DM, respectively. Mixed duckweeds as a component of a concentrate mixture were eaten by the cattle at the rate of 10% of their live weights. It may be concluded that the dry matter and crude protein of the available duckweeds wee highly degradable in the rumen and may be fed to cattle mixing with concentrates. For the effective utilization of duck weeds as cattle feed their effect on the rumen digestion kinetics of a roughage diet need to be studied carefully.

• 대한미생물학회지
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• 제35권4호
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• pp.273-282
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• 2000

dTDP-rhamnose provides L-rhamnose to the bridge-like structure between mycolyl arabinogalactan and peptidoglycan of the mycobacterial cell wall. dTDP-rhamnose is composed of glucose-l-phosphate and dTTP by four enzymes encoded by rmlA-D. To determine the region(s) of RmlC protein essential for its dTDP-4-keto-6-deoxyglucose epimerase activity, we overexpressed both whole (202 amino acids) and three different truncated (N-terminal 106 or 150 or C-terminal 97 amino acids) RmlC proteins of Mycobacterium tuberculosis. The RmlC enzyme activity in the soluble lysates of ${\Delta}rmlC$ E. coli strain $S{\Phi}874$ (DE3 PlysS) expressing the wild type or truncated rmlC genes was initially analyzed by three sequential reactions from dTDP-glucose to dTDP-rhamnose in the presence of purified RmlB and RmlD. All three soluble lysates containing the truncated RmlC proteins showed no enzyme activity, while that containing the wild type RmlC was active. This wild type RmlC was then overexpressed and purified. The incubation of the purified RmlC enzyme so obtained with dTDP-4-keto-6-deoxyglucose resulted in the conversion of dTDP-4-keto-rhamnose. The results show that the truncated regions of the RmlC protein are important for the RmlC enzyme activity in M. tuberculosis.

• Advances in nano research
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• 제10권2호
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• pp.191-210
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• 2021

To compare the antifungal effect of two nanomaterials (NMs), nanoparticles of zinc oxide were synthesized by a chemical route and zinc oxide-based nanobiohybrids were obtained using green synthesis in an extract of garlic (Allium sativum). The techniques of X-Ray Diffraction (XRD), Infrared (IR) and Ultraviolet Visible (UV-Vis) absorption spectroscopies and Scanning (SEM) and Transmission Electron Microscopies (TEM) were used to determine the characteristics of the nanomaterials synthesized. The results showed that the samples obtained were of nanometric size (< 100 nm). To compare their antifungal capacity, their effect on Cercospora sp. was evaluated. Test results showed that both nanomaterials had an antifungal capacity. The nanobiohybrids (green route) gave an inhibition of fungal growth of ~72.4% while with the ZnO-NPs (chemical route), inhibition was ~87.1%. Microstructural studies using High Resolution Optical Microscopy (HROM) and ultra-structural analysis using TEM carried out on the treated strains demonstrated the effect of the nanofungicides on the vegetative and reproductive structures, as well as on their cell wall. To account for the antifungal effect presented by ZnO-NPs and ZnO nanobiohybrids on the fungi tested, effects reported in the literature related to the action of nanomaterials on biological entities were considered. Specifically, we discuss the electrical interaction of the ZnO-NPs with the cell membrane and the biomolecules (proteins) present in the fungi, taking into account the n-type nature of the ZnO semiconductor and the electrical behavior of the fungal cell membrane and that of the proteins that make up the protein crown.

• Journal of Microbiology and Biotechnology
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• 제33권7호
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• pp.964-972
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• 2023

Bacteriophage endolysins are peptidoglycan hydrolases composed of cell binding domain (CBD) and an enzymatically active domain. A phage endolysin CBD can be used for detecting bacteria owing to its high specificity and sensitivity toward the bacterial cell wall. We aimed to develop a method for detection of Enterococcus faecalis using an endolysin CBD. The gene encoding the CBD of ECP3 phage endolysin was cloned into the Escherichia coli expression vector pET21a. A recombinant protein with a C-terminal 6-His-tag (CBD) was expressed and purified using a His-trap column. CBD was adsorbed onto epoxy magnetic beads (eMBs). The bacterial species specificity and sensitivity of bacterial binding to CBD-eMB complexes were determined using the bacterial colony counting from the magnetic separations after the binding reaction between bacteria and CBD-eMB complexes. E. faecalis could bind to CBD-eMB complexes, but other bacteria (such as Enterococcus faecium, Staphylococcus aureus, Escherichia coli, Acinetobacter baumannii, Streptococcus mutans, and Porphyromonas gingivalis) could not. E. faecalis cells were fixed onto CBD-eMB complexes within 1 h, and >78% of viable E. faecalis cells were recovered. The E. faecalis recovery ratio was not affected by the other bacterial species. The detection limit of the CBD-eMB complex for E. faecalis was >17 CFU/ml. We developed a simple method for the specific detection of E. faecalis using bacteriophage endolysin CBD and MBs. This is the first study to determine that the C-terminal region of ECP3 phage endolysin is a highly specific binding site for E. faecalis among other bacterial species.

• 한국초지조사료학회지
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• 제5권3호
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• pp.180-186
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• 1985

본(本) 시험(試驗)은 옥수수와 sorghum식물(植物)에서 생육시기(生育時期) 및 환경온도(環境溫度)가 Weender성분(成分) 및 Net Energy Lactation 에 미치는 영향(影響)을 구명(究明)코자 옥수수의 Biizzard와 sorghum hybrids와 Pioneer931 및 Sioux를 공시품종(供試品種)으로 하여 뮌헨대학교초지연구소(大學校草地硏究所)에서 포장(圃場) 및 phytotron시험(試驗)으로 실시(實施)하였다. Phytotron의 주(晝)/야간(夜間) 실내온도(室內溫度)는 30/25, 25/20, 28/18 및 $18/8^{\circ}C$로 하였으며 일조(日照)는 35,000Lux로 13시간(時間) 조사(照謝)하였다. 1978-'81년간(年間) 얻어진 결과(結果)를 요약(要約)하면 다음과 같다. 1. 옥수수 및 sorghum의 crude protein은 3 엽기유식물(葉期幼植物)의 경우 각각(各各) 31.4% 및 33.9%에 이르나 생육(生育)이 진전(進展)됨에 따라 급속도(急速度)로 하락(下落)되어 출수기(出穗期)에는 9.0%(옥수수) 및 10.4% (sorghum)로 감소(減少)된다. 식물체내의 protein축적(蓄積)과 LWR 및 LAR간(間)에는 높은 정(正)(+)의 상관(相關)이 있다.($P{\leq}0.1%$). 2. 조섬유(粗纖維)의 합성(合成) 및 축적(蓄積)은 유수형성기(幼穗形成期)에서 지엽출현기(止葉出現期)에 본격적(本格的)으로 이루어지며 그 함량(含量)은 출수기(出穗期)에 각각(各各) 옥수수 28.4% 및 Sorghum 31.5%로 최고농도수집(最高濃度水準)에 달한다. 그러나 옥수수의 경우 조섬유(粗纖維) 함량(含量)은 종자(種子)가 성숙(成熟)됨에 따라 다시 감소(減少)되어 황숙기(黃熟期)에는 19.5%까지 감소(減少)되는데 반(反)해 sorghum 식물(植物)에서는 계속적(繼續的)으로 증가(增加)하는 경향(傾向)이 있다. 3. NEL 함량(含量)은 3 엽기(葉期)의 유식물(幼植物)에서 옥수수 5.98MJ 및 sorghum 5.64MJ/kg으로 높은 편이나 생육(生育)이 진행(進行)되는 동안 감소(減少)되어 유수(幼穗)가 형성(形成)되는 6-8 엽기(葉期)에 각각(各各) 5.82MJ(옥수수) 및 5.46MJ/kg(sorghum)으로 최저수준(最低水準)을 나타낸다. 옥수수의 NEL 함량(含量)은 그후 종자(種子)가 성숙(成熟)됨에 따라 6.70MJ(乳熟期) 및 6.94MJ/kg(完熟期)까지 증가(增加)되는데 비해 sorghum에서는 증가폭(增加幅)이 완만하여 계속(繼續) 낮은 수준(水準)을 유지(維持)한다. 4. 옥수수 및 sorghum식물(植物)의 NEL가치(價値)는 fruetosan, mono- 및 disaccharose등 non-structural carbo-hydrate의 합성(合成) 및 축적형태(蓄積形態)에 의(依)해 큰 영향(影響)을 받으면 NEL함량(含量)과 cell-wall constituents간(間)에는 부(負)(-)의 상관(相關)이 있다.($P{\leq}0.01%$). 5. NEL 및 starch value 환경온도(環境溫度)가 상승(上昇)됨에 따라 감소(減少)된다. 4 엽기(葉期) sorghum식물(植物)의 환경온도(環境溫度)를 달리 하였을 때 NEL가치(價値)는 각각(各各) 4.87MJ($30/25^{\circ}C$), 5.46MJ($25/20^{\circ}C$) 및 5.81MJ/kg($18/8^{\circ}C$)로 변(變)하여 고온(高溫)에서 net energy lactation 축적(蓄積)이 크게 감소(減少)되었다.

• 한국식품영양과학회지
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• 제29권3호
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• pp.401-406
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• 2000

가공식품의 개발에 있어서 식품의 맛과 더불어 저장성, 열안정성 및 색조유지는 소비자의 기호도에 중요한 영향을 미친다. B.subtilis EK11 유래의 PPase는 식물조직 중엽부의 주성분인 불용성 protopectin을 분해하여 단세포화하는 효소이다. PPase를 참다래에 작용시켜 참다래 고유의 세포 속에 함유되어 있는 세포내 성분들의 파손 없이 단세포를 유리하였다. 참다래 조직으로부터 제조된 단세포화물의 착즙 후 관찰된 회수율과 잔사율은 각각 82%와 18%로써, 기계적 마쇄물에서의 60%와 40%에 비하여 높은 회수율과 낮은 잔사율을 나타내었다. 총당, 환원단, sucrose, 조단백질, 조지방 및 조섬유에 대해 함량 변화는 큰 차이가 없었으며, 이는 단세포 처리에 의하여 이들 성분이 안전하게 유지됨을 알 수 있었다. PPase로 처리시 가장 불안정한 비타민 C가 1일 경과 후에도 95% 이상이 보존되는 것으로 보아 단세포에 의한 일반적인 구성성분이 안정하게 유지 보호됨을 알 수 있다. PPase로 처리된 참다래 단세포물을 $4^{\circ}C$에서 6일간 저장하며 색조를 관찰한 결과, 기계적 마쇄물에서는 변색이 일어났으나 단세포화물에서는 뚜렷한 색조의 차이가 없었다. 또한 PPase로 처리한 참다래 단세포물을 $100^{\circ}C$에서 60분간 열처리한 후 관찰한 결과, 기계적 마쇄물에서는 짧은 처리에도 극심한 변화를 보였으나 단세포화물에서는 변화가 보이지 않아 높은 열안정성을 나타내었다. 따라서 PPase는 참다래의 단세포화물에 응용 가능하여 참다래 음료제조 및 원료 보존에 유용하게 이용할 수 있을 것으로 여겨 진다. 또한 PPase을 이용한 참다래의 효율성 제고와 고부가가치의 기능성 식품제조에 이용될 수 있음을 의미한다.