• Title/Summary/Keyword: fluorescence control

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Use of Human Serum Albumin Fusion Tags for Recombinant Protein Secretory Expression in the Methylotrophic Yeast Hansenula polymorpha (메탄올 자화효모 Hansenula polymorpha에서의 재조합 단백질 분비발현을 위한 인체 혈청 알부민 융합단편의 활용)

  • Song, Ji-Hye;Hwang, Dong Hyeon;Oh, Doo-Byoung;Rhee, Sang Ki;Kwon, Ohsuk
    • Microbiology and Biotechnology Letters
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    • v.41 no.1
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    • pp.17-25
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    • 2013
  • The thermotolerant methylotrophic yeast Hansenula polymorpha is an attractive model organism for various fundamental studies, such as the genetic control of enzymes involved in methanol metabolism, peroxisome biogenesis, nitrate assimilation, and resistance to heavy metals and oxidative stresses. In addition, H. polymorpha has been highlighted as a promising recombinant protein expression host, especially due to the availability of strong and tightly regulatable promoters. In this study, we investigated the possibility of employing human serum albumin (HSA) as the fusion tag for the secretory expression of heterologous proteins in H. polymorpha. A set of four expression cassettes, which contained the methanol oxidase (MOX) promoter, translational HSA fusion tag, and the terminator of MOX, were constructed. The expression cassettes were also designed to contain sequences for accessory elements including His8-tag, $2{\times}(Gly_4Ser_1)$ linkers, tobacco etch virus protease recognition sites (Tev), multi-cloning sites, and strep-tags. To determine the effects of the size of the HSA fusion tag on the secretory expression of the target protein, each cassette contained the HSA gene fragment truncated at a specific position based on its domain structure. By using the Green fluorescence protein gene as the reporter, the properties of each expression cassette were compared in various conditions. Our results suggest that the translational HSA fusion tag is an efficient tool for the secretory expression of recombinant proteins in H. polymorpha.

Evaluation of Odors and Odorous Compounds from Liquid Animal Manure Treated with Different Methods and Their Application to Soils (액상 가축분뇨의 처리 및 토양환원에 따른 악취 및 악취물질의 평가)

  • 고한종;최홍림;김기연;이용기;김치년
    • Journal of Animal Science and Technology
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    • v.48 no.3
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    • pp.453-466
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    • 2006
  • To comply with stricter regulations provoked by increasing odor nuisance, it is imperative to practice effective odor control for sustainable livestock production. This study was conducted to assess odor and odorous compounds emitted from liquid animal manure with different treatment methods such as Fresh Manure(without treatment, FM), Anaerobic Digestion(AD) and Thermophilic Aerobic Digestion(TAD) and their application to soil. Air samples were collected at the headspace of liquid manure, upland and paddy soil, and analyzed for odor intensity and offensiveness using an olfactometry; odor concentration index using odor analyser; nitrogen-containing compound such as ammonia(NH3) using fluorescence method; and sulfur containing compounds such as hydrogen sulfide(H2S), methyl mercaptan(MeSH), dimethyl sulfide(DMS) and dimethyl disulfide(DMDS) using gas chromatography-pulsed flame photometric detector, respectively. Odor intensity, offensiveness and concentration index from TAD liquid manure was statistically lower than those from FM and AD(p<0.01). Mean concentrations of H2S, MeSH, DMS, DMDS and NH3 were 65.93ppb, 18.55ppb, 5.26ppb, 0.33ppb and 10.57ppm for liquid manure with AD; and 5.15ppb, 0.97ppb, 0.80ppb, 0.56ppb and 1.34ppm for liquid manure with TAD, respectively. More than 60% of malodorous compounds related to nitrogen and sulfur were removed by heterotrophic microorganisms during TAD treatment. When liquid manure was applied onto upland and paddy soil, NH3 removal efficiencies ranged from 51 to 94% and 22 to 91% for AD and TAD liquid manure, respectively. The above results show that liquid manure with TAD is superior to AD and FM with respect to the odor reduction and odor problem caused by land applied liquid manure is directly related to the degree of odor generated by the manure treatment method.

Effects of Hydrogen Peroxide on Germination and Early Growth of Sorghum (Sorghum bicolor) (과산화수소 처리가 수수의 발아 및 초기 생장에 미치는 효과)

  • Shim, Doobo;Song, Ki Eun;Park, Chan Young;Jeon, Seung Ho;Hwang, Jung Gyu;Kang, Eun-ju;Kim, Jong Cheol;Shim, Sangin
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.63 no.2
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    • pp.140-148
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    • 2018
  • As the global warming causing desertification increase, there is growing concern about damage of crops. It was to investigate how the treatment with hydrogen peroxide before leaf development affects the growth and yield of sorghum for minimizing a damage of crops to drought. The germination experiment was conducted at alternating temperature of $25^{\circ}C/20^{\circ}C$(12 hr/12 hr) under water stress condition of 0 ~ -0.20 MPa adjusted with PEG solution containing 0 and 10 mM $H_2O_2$. In order to know the effect of foliar application of hydrogen peroxide on the growth of sorghum, 10 mM hydrogen peroxide was treated to leaves at 3-leaf stage of sorghum growing in greenhouse conditions. Seed germination rate was increased by 20% in hydrogen peroxide treatment as compared to the Control. under water stress conditions (-0.15 ~ -0.20 MPa). The length of seedlings was also on the rise by the hydrogen peroxide treatment. In the greenhouse pot experiment, the morphological characteristics (plant height, stem diameter, leaf length, and leaf number) and physiological characteristics (chlorophyll content, chlorophyll fluorescence (Fv/Fm), stomatal conductance) were higher in the plants treated with hydrogen peroxide under the drought stress condition than those of plants of $H_2O$ treatment. Experiment conducted with the soil moisture gradient system showed that the foliar application of hydrogen peroxide increased photosynthetic ability of sorghum plant with respect to SPAD value and stomatal conductance and rooting capacity (root weight and root length) under drought condition. Generally, hydrogen peroxide treatment in sorghum increased the tolerance to drought stress and maintained better growth due to ameliorating oxidative stress.

Effects of Differentiated Temperature Based on Growing Season Temperature on Growth and Physiological Response in Chinese Cabbage 'Chunkwang' (고랭지 여름배추 주산지의 기온을 기준으로 한 수준별 온도가 배추 '춘광'의 생육 및 생리반응에 미치는 영향)

  • Son, In-Chang;Moon, Kyung Hwan;Song, Eun Young;Oh, Soonja;Seo, Hyeongho;Moon, Young Eel;Yang, Jinyoung
    • Korean Journal of Agricultural and Forest Meteorology
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    • v.17 no.3
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    • pp.254-260
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    • 2015
  • Changes of the growth, quality and physiological response of Chinese cabbage cv 'Chunkwang' in response to five different temperature treatments based on climate change scenario were investigated during the growing season. The treatments consisted of normal year temperature $-2.0^{\circ}C$ (I), normal year temperature (II; Control group), normal year temperature $+2.0^{\circ}C$ (III), normal year temperature $+4.0^{\circ}C$ (IV), and normal year temperature $+6.0^{\circ}C$ (V). Regarding fresh weight, number of leaves, and leaf area were high in group IV, and V before the head formation stage, but it has decreased during the later growth period. Rate of frangibleness sympton was the highest in group V as 85.7%, and it was decreased in group IV (64.3%), group III (28.6%), group II (14.3%), and group I (7.1%). Regarding photosynthetic rate, group III, IV, and V showed relatively high photosynthetic rate at 20 DAP but it was reduced dramatically during the later growth period. Transpiration and stomatal conductance showed the similar trend with the photosynthetic rate. When comparing the chlorophyll fluorescence reaction of each treatment group at 50 DAP, Fv/Fm in group I was highest as 8.04 among all treatment groups and the lowest in group IV as 7.15.

Studies on the Control of the Pine Bark Beetle (Myelophilus pinipedera LINNE) (I) -Analysis of the carotenoids in important pine species in Korea- (소나무좀 방제(防除)에 관한 연구(硏究)(제1보(第1報)) -한국산(韓國産) 중요(重要)소나무류(類) 잎에 함유(含有)되어 있는 Carotenoid 분석(分析)-)

  • Kim, Young Ho
    • Journal of Korean Society of Forest Science
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    • v.27 no.1
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    • pp.15-21
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    • 1975
  • According to Yoshihisa Kuroki's report that the total amount of carotenoid was more in the susceptible to pine bark beetle, than in the resistant ones, carotenoids were extracted from needle leaves of one year old seedlings of Pinus thunbergii, Pinus desiflora, Pinus rigida, Pinus koraiensis, Pinus rigitaeda and Pinus taeda which are all important pines in Korea, to find their resistant ratio to the insect. The carotenoids were analyzed and compared using the spectra of them by spectrophotometer. The results were as follows: 1. The visible absorption spectra of carotenoids in those pine trees were proved to be very similar. 2. The total amount of carotenoids in needles differed with the tree species and the contents were arranged in decreasing order P. koraiensis>P. rigida>P. thunbergii>P. rigitaeda>P. taeda, it seemed that there was corelation between the cartenoid content and the extent of harm caused by the pine bark beetles except P. koraiensis. 3. But carotenoids were contained in Robinia pseudoacacia, Castanea crenata, Chamaecyparis obtusa and Cedrus deodra leaves too. 4. The total amounts of carotenoids in these pines of 9 species were arranged in decreasing order Robinia pseudoacacia>Pinus koraiensis>Pinus rigida>Pinus thunbergii>Castanea crenata>Pinus rigitaeda>Pinus taeda>Chamaecyparis obtusa>Cedrus deodara. Therefore, it was proved that there was no correlation between carotenoid cotent and extent of resistance to the insect. 5. In the thin-layer chromatography of these carotenoids, 13 kinds of components in P. densiftora, P. koraiensis and P. rigida and 12 kinds of spots in other pines, were detected respectively, under ultra-violet fluorescent lamp $3,600{\AA}$ and $2,537{\AA}$. 6. The eighth spots from the bottom in P. densiflora, P. koraiensis and P. rigida were not found in other pines and other 4 species (Robinia pseudoacacia, Castanea crenata, Chamacyparis obtusa, Cedrus deodra). Especially the spot in P. densiflora fluoresced strong cobalt blue-fluorescence under ultra-violet fluorescent lamp $2,537{\AA}$.

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Transfer of Isolated Mitochondria to Bovine Oocytes by Microinjection (미세주입을 이용한 난자로의 분리된 미토콘드리아 전달)

  • Baek, Sang-Ki;Byun, June-Ho;Kim, Bo Gyu;Lee, A ram;Cho, Young-Soo;Kim, Ik-Sung;Seo, Gang-Mi;Chung, Se-Kyo;Lee, Joon-Hee;Woo, Dong Kyun
    • Journal of Life Science
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    • v.27 no.12
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    • pp.1445-1451
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    • 2017
  • Mitochondria play a central role in energy generation by using electron transport coupled with oxidative phosphorylation. They also participate in other important cellular functions including metabolism, apoptosis, signaling, and reactive oxygen species production. Therefore, mitochondrial dysfunction is known to contribute to a variety of human diseases. Furthermore, there are various inherited diseases of energy metabolism due to mitochondrial DNA (mtDNA) mutations. Unfortunately, therapeutic options for these inherited mtDNA diseases are extremely limited. In this regard, mitochondrial replacement techniques are taking on increased importance in developing a clinical approach to inherited mtDNA diseases. In this study, green fluorescence protein (GFP)-tagged mitochondria were isolated by differential centrifugation from a mammalian cell line. Using microinjection technique, the isolated GFP-tagged mitochondria were then transferred to bovine oocytes that were triggered for early development. During the early developmental period from bovine oocytes to blastocysts, the transferred mitochondria were observed using fluorescent microscopy. The microinjected mitochondria were dispersed rapidly into the cytoplasm of oocytes and were passed down to subsequent cells of 2-cell, 4-cell, 8-cell, morula, and blastocyst stages. Together, these results demonstrate a successful in vitro transfer of isolated mitochondria to oocytes and provide a model for mitochondrial replacement implicated in inherited mtDNA diseases and animal cloning.

Trial for Drug Susceptibility Testing of Mycobacterium tuberculosis with Live and Dead Cell Differentiation (세포 염색 방법을 이용한 결핵균 감수성 검사법)

  • Ryu, Sung-Weon;Kim, Hyun-Ho;Bang, Mun-Nam;Park, Young-Kil;Park, Sue-Nie;Shim, Young-Soo;Kang, Seongman;Bai, Gill-Han
    • Tuberculosis and Respiratory Diseases
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    • v.56 no.3
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    • pp.261-268
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    • 2004
  • Background : The resurgence of tuberculosis and outbreaks of multidrug resistant (MDR) tuberculosis have increased the emphasis for the development of new susceptibility testing of the Mycobacterium tuberculosis for the effective treatment and control of the disease. Conventional drug susceptibility testings, such as those using egg-based or agar-based media have some limits, such as the time required and difficulties in determining critical inhibitory concentrations, but these are still being used in many diagnostic laboratories because of no better lternatives, considering cost and accuracy. To overcome these limits, a rapid and simple method for new susceptibility testing, using live and dead assays, was applied for a bacterial cell viability assay to distinguish dead from live bacterial cells based on two-color fluorescence. Materials and Methods Strains : Forty strains were used in this study, 20 susceptible to all antituberculosis drugs and the other 20 resistant to the four first line antituberculosis drugs isoniazid, rifampicin, streptomycin and ethambutol. Antibiotics : The four antibiotics were dissolved in 7H9 broth to make the following solutions: $0.1{\mu}g\;isoniazid(INH)/m{\ell}$, $0.4{\mu}g\;rifampicin(RMP)/m{\ell}$, $4.0{\mu}g\;streptomycin(SM)/m{\ell}$ and $4.0{\mu}g\;ethambutol(EMB)/m{\ell}$. Results : Live and dead Mycobacterium tuberculosis cells fluoresced green and red with the acridin (Syto 9) and propidium treatments, respectively. These results are very well accorded with conventional drug susceptibility testing by proportional method on Lowensen-Jensen media (L-J) containing 4 drugs (INH, RMP, EMB and SM), showing a 93.7 % accordance rate in susceptible strains and 95% in resistant strains. Conclusion : The results of the drug susceptibility testing using the live and dead bacterial cell assay showed high accordance rates compared with the conventional proportion method on L-J. This finding suggests that the live and dead bacterial cell assay can be used as an alternative to conventional drug susceptibility testing for M. tuberculosis strains.

Prelaunch Study of Validation for the Geostationary Ocean Color Imager (GOCI) (정지궤도 해색탑재체(GOCI) 자료 검정을 위한 사전연구)

  • Ryu, Joo-Hyung;Moon, Jeong-Eon;Son, Young-Baek;Cho, Seong-Ick;Min, Jee-Eun;Yang, Chan-Su;Ahn, Yu-Hwan;Shim, Jae-Seol
    • Korean Journal of Remote Sensing
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    • v.26 no.2
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    • pp.251-262
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    • 2010
  • In order to provide quantitative control of the standard products of Geostationary Ocean Color Imager (GOCI), on-board radiometric correction, atmospheric correction, and bio-optical algorithm are obtained continuously by comprehensive and consistent calibration and validation procedures. The calibration/validation for radiometric, atmospheric, and bio-optical data of GOCI uses temperature, salinity, ocean optics, fluorescence, and turbidity data sets from buoy and platform systems, and periodic oceanic environmental data. For calibration and validation of GOCI, we compared radiometric data between in-situ measurement and HyperSAS data installed in the Ieodo ocean research station, and between HyperSAS and SeaWiFS radiance. HyperSAS data were slightly different in in-situ radiance and irradiance, but they did not have spectral shift in absorption bands. Although all radiance bands measured between HyperSAS and SeaWiFS had an average 25% error, the 11% absolute error was relatively lower when atmospheric correction bands were omitted. This error is related to the SeaWiFS standard atmospheric correction process. We have to consider and improve this error rate for calibration and validation of GOCI. A reference target site around Dokdo Island was used for studying calibration and validation of GOCI. In-situ ocean- and bio-optical data were collected during August and October, 2009. Reflectance spectra around Dokdo Island showed optical characteristic of Case-1 Water. Absorption spectra of chlorophyll, suspended matter, and dissolved organic matter also showed their spectral characteristics. MODIS Aqua-derived chlorophyll-a concentration was well correlated with in-situ fluorometer value, which installed in Dokdo buoy. As we strive to solv the problems of radiometric, atmospheric, and bio-optical correction, it is important to be able to progress and improve the future quality of calibration and validation of GOCI.

Effects of Increasing Air Temperatures and CO2 Concentrations on Herbicide Efficacy of Acalypha australis and Phytotoxicity of Soybean Crops (대기온도와 CO2 농도 증가에 따른 우점잡초 깨풀의 제초제 약효 및 콩 약해 변화)

  • Hyo-Jin Lee;Hyun-Hwa Park;Ye-Geon Kim;Do-Jin Lee;Yong-In Kuk
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.68 no.3
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    • pp.121-133
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    • 2023
  • The purpose of this study was to improve weed management systems under varying carbon dioxide concentrations and temperatures by evaluating the growth of Acalypha australis and observing the efficacy of four foliar and four soil herbicides, as well as measuring phytotoxicity in soybean crops treated with these herbicides. In both growth chamber and greenhouse conditions, plant height and shoot fresh weight of Acalypha australis increased as temperature increased. The variable to maximum fluorescence ratio (Fv/Fm), relative electron transport rate (ETR), plant height, leaf area, and shoot fresh weight of Acalypha australis were higher at carbon dioxide concentrations of 800 ppm than at 400 ppm. The efficacy of a foliar herbicide, glufosinate, on Acalypha australis was lower at 30℃ than at 20℃ and 25℃ in the growth chamber condition and was also lower at 29℃ than at 21℃ and 25℃ in greenhouse conditions. In contrast, mecoprop efficacy on Acalypha australis was lower at 20℃ and 25℃ than at 30℃ in growth chamber conditions and lower at 21℃ and 25℃ than at 29℃ in greenhouse conditions. Glyphosate efficacy was lower at 21℃ than at 25℃ and 29℃ under greenhouse conditions. With soil herbicides, metolachlor and ethalfluraline, efficacies were higher at relatively high temperatures under both growth chamber and greenhouse conditions. However, in the case of linuron, the difference in efficacy was not observed under varying temperatures in both growth chamber and greenhouse conditions. When ¼ of the recommended glyphosate rates were applied to Acalypha australis, efficacy was lower under 800 ppm carbon dioxide concentrations than under 400 ppm. In contrast, when ¼ of the recommended rate of bentazone was applied to Acalypha australis, efficacy was higher under 800 ppm carbon dioxide concentrations than under 400 ppm. Despite application rates, glufosinate efficacy differed insignificantly under different carbon dioxide concentrations. When applied at ¼ of the recommended rate, the efficacy of ethalfuralin was higher under 800 ppm carbon dioxide concentrations than under 400 ppm. However, efficacies of other herbicides were not different despite varying carbon dioxide concentrations. Soybean phytotoxicity in crops treated with the recommended rate and twice the recommended rate of soil herbicides was not significantly different regardless of temperature and carbon dioxide concentrations. Overall, weed efficacy of some herbicides decreased in response to different temperatures and carbon dioxide concentrations. Therefore, new weed management methods are required to ensure high rates of weed control in conditions affected by climate change.