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Development of System-Wide Functional Analysis Platform for Pathogenicity Genes in Magnaporthe oryzae

  • Park, Sook-Young;Choi, Jaehyuk;Choi, Jaeyoung;Kim, Seongbeom;Jeon, Jongbum;Kwon, Seomun;Lee, Dayoung;Huh, Aram;Shin, Miho;Jung, Kyungyoung;Jeon, Junhyun;Kang, Chang Hyun;Kang, Seogchan;Lee, Yong-Hwan
    • 한국균학회소식:학술대회논문집
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    • 2014.10a
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    • pp.9-9
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    • 2014
  • Null mutants generated by targeted gene replacement are frequently used to reveal function of the genes in fungi. However, targeted gene deletions may be difficult to obtain or it may not be applicable, such as in the case of redundant or lethal genes. Constitutive expression system could be an alternative to avoid these difficulties and to provide new platform in fungal functional genomics research. Here we developed a novel platform for functional analysis genes in Magnaporthe oryzae by constitutive expression under a strong promoter. Employing a binary vector (pGOF1), carrying $EF1{\beta}$ promoter, we generated a total of 4,432 transformants by Agrobacterium tumefaciens-mediated transformation. We have analyzed a subset of 54 transformants that have the vector inserted in the promoter region of individual genes, at distances ranging from 44 to 1,479 bp. These transformants showed increased transcript levels of the genes that are found immediately adjacent to the vector, compared to those of wild type. Ten transformants showed higher levels of expression relative to the wild type not only in mycelial stage but also during infection-related development. Two transformants that T-DNA was inserted in the promotor regions of putative lethal genes, MoRPT4 and MoDBP5, showed decreased conidiation and pathogenicity, respectively. We also characterized two transformants that T-DNA was inserted in functionally redundant genes encoding alpha-glucosidase and alpha-mannosidase. These transformants also showed decreased mycelial growth and pathogenicity, implying successful application of this platform in functional analysis of the genes. Our data also demonstrated that comparative phenotypic analysis under over-expression and suppression of gene expression could prove a highly efficient system for functional analysis of the genes. Our over-expressed transformants library would be a valuable resource for functional characterization of the redundant or lethal genes in M. oryzae and this system may be applicable in other fungi.

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Treatment of Canine Cervical and Lumbar Disc Disease by Injection-Acupuncture (개 경부 및 요부 디스크의 수침치료)

  • Kim Duck-Hwan;Liu Jian-Zhu;Lee Young-Won;Song Kun-Ho;Kang Sang-Kyu;Choi Ho-Jung;Seo Kang-Moon;Choi Seok-Hwa;Nam Tchi-Chou;Rogers Phil A.M.
    • Journal of Veterinary Clinics
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    • v.23 no.1
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    • pp.65-68
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    • 2006
  • Two cases of canine intervertebral disc disease (IVDD) were treated twice a week with injection-acupuncture (injection-AP) using 0.1 ml/acupoint. In case 1, a paralysed dog with cervical intervertebral disc disease (C3-C4), was submitted to injection-AP with dexamethasone (1 mg/ml) at session 1-2 and thiamine (25 mg/ml) at session 3-5. Injected acupoints included GV-16, GB-20, BL-10, LU-7, LI-4 and SI-6. The acupoints GV-6, GV-20 and SP-6 were added at session 2-5. Trigger point (TP) therapy with 0.2 ml of 2% lidocaine was used in session 3-5 at TP in the infraspinatus and triceps muscles. Treatment was stopped when paralysis disappeared after 5 injection-AP treatments; there were no recurrent symptoms in the follow-up period of 5 months. In case 2, an ataxic dog with lumbar IVDD (L1-L2), was submitted to injection-AP with dexamethasone at session 1 and 2 and thiamine at session 3-4. Injected acupoints included GV-6 as the main point, ST-36, GB-30, ST-40, GB-34, ST-41 and BL-40. TP therapy with 0.2 ml of 2% lidocaine was used at TP in the ileocostorum lumborum and quadriceps muscles. Treatment was stopped when ataxia disappeared after 4 treatments; there were no recurrent symptoms In the fallow-up period of 5 months. Injection-AP using dexamethasone and thiamine, combined with TP therapy using lidocaine, effectively alleviate the symptoms of canine cervical and lumbar IVDD.

Studies on the postnatal development of the Leydig cell in rabbits (토끼에서 출생 후 고환간질세포의 발생에 관한 연구)

  • Tae, Hyun-Jin;Park, Young-Jae;Kang, Hyung-Sub;Kim, Nam-Soo;Park, Sang-Youel;Yang, Hong-Hyun;Ahn, Dong-Choon;Kim, In-Shik
    • Korean Journal of Veterinary Research
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    • v.45 no.3
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    • pp.325-334
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    • 2005
  • Changes in the rabbit Leydig cell from birth to adulthood were studied in New Zealand white rabbits of 1, 7, 21, 35, 49, 70, 105, 147, 196, and 252 days (n = 8 rabbits per group) of age. The objectives of this study were to understand the fate of the fetal Leydig cells, to determine the changes in serum testosterone levels, and leutenizing hormone-stimulated testosterone production per testis in vitro, and to quantify adult Leydig cells by number and average volume with age. Testes of rabbits were fixed by whole body perfusion using a fixative containing 2.5% glutaraldehyde in cacodylate buffer, processed and embedded in Epon-araldite. Using $1{\mu}m$ sections stained with methylene blue-azure II, qualitative and quantitative (stereological) morphological studies were performed. Testosterone levels in the incubation medium of luteinizing hormone-stimulated (100 ng/ml) testosterone secretion per testis in vitro, and in serum were determined by radioimmunoassay. The average volume of a testis of 1-day-old rabbits was determined as $0.0073cm^3$ and the parameter increased linearly from birth to 252 days ($3.93cm^3$). The volume density of the seminiferous tubules increased with age from 33.76% at day 1 to 88.2% at day 252. The volume density of the interstitium represents 66.24% of the testicular parenchyma at day 1. This proportion progressively diminished during development to reach a value of 11.8% at day 252. The volume density of Leydig cells increased almost linearly from birth (0.001%) to 252 days (2.62%). Leydig cell mass per testis increases from 0.0012 mg to 0.25 mg between days 1 and 35, from 2.66 mg to 44.3 mg between days 49 and 105 and from 65.42 mg and 102.9 mg between days 147 and 252. The absolute numbers of adult Leydig cells per testis increased linearly from birth to 252 days. The average volume of adult Leydig cell on days 1, 7, 21 and 35 was not significantly different; a gradual and continued increase was observed thereafter, reaching a 3-fold increase at 196 and 252 days. Serum testosterone concentrations were not significantly different at day 1 compared days 7, 21, 35. Significant increases were observed at days 49 and 70. Values at days 70 and 105 and days 147, 196, and 252 were not significantly different. LH-stimulated testosterone production per testis in vitro was significantly different at day 1 compared days 7, 21, 35. Significant increases were observed at days 49 and 70. Hormonal values at days 105, 147, 196, and 252 were not significantly different. These data suggested Leydig cell developmental phase can be classified: a neonatal phase (1-7 days), a prepubertal phase (14-49 days) and an adult phase (70-252 days). Immature and mature adult Leydig cells, initially detected at days 7 and 49, respectively, and mature adult Leydig cells were abundant Leydig cell type according to the number and absolute volume per testis form day 49 onwards.

Ecological Study on the Seed Germination and Emergence of Overwintered Stump of Bulrush(Scirpus juncoides Roxb.) (올챙고랭이(Scirpus juncoides Roxb.) 종자발아(種子發芽) 및 월동(越冬) 주기부(株基部)의 출아생태(出芽生態)에 관한 연구(硏究))

  • Guh, J.O.;Huh, S.M.
    • Korean Journal of Weed Science
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    • v.6 no.2
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    • pp.110-123
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    • 1986
  • To know the ecological pattern of bulrush (Scirpus juncoides Roxb.) seeds and overwintreed stumps in germination and sprouting responses as affected by different temperature (7 trt.), light intensity (5 trt.), shading intensity (S trt.), light quality (specturm spectrum; 6 trt.), soil acidity (7 trt.), stump sizea (weight base; 5 trt.), and molding depth (6trt.), respectively, this serial studies were conducted by use of growth chamber, incubator, Wagner pot and petri-dish. Most efficient treatment was obtained from 25-$35^{\circ}C$ temperature, higher light intensity in 2-11 klux range, 95% shading intensity, clear and yellow film for seeds/clear and blue film for stumps, soil pH 5.53, 3-4g stump weight, 0-5% wxygen concentration, 1 ㎝ flooding depth for seeds, and 1-1.5cm molding depth for seeds/0.5-1.0 cm molding depth for stumps, respectively, among others.

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Mechanism of Growth Inhibition in Herbicide-Resistant Transgenic Rice Overexpressing Protoporphyrinogen Oxidase (Protox) Gene (Protoporphyrinogen Oxidase (Protox) 유전자 과다발현 제초제 저항성 형질전환 벼의 생육저해 기작)

  • Kuk, Yong-In;Shin, Ji-San;Yun, Young-Beom;Kwon, Oh-Do
    • Korean Journal of Weed Science
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    • v.30 no.2
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    • pp.122-134
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    • 2010
  • We investigated the levels of resistance and accumulation of terapyrroles, reactive oxygen species, lipid peroxidation, and antioxidative enzymes for reasons of growth reduction in herbicide-transgenic rice overexpressing Myxococcus xanthus, Arabidopsis thaliana, and human protoporphyrinogen oxidase (Protox) genes. The transgenic rice overexpressing M. xanthus (MX, MX1, PX), A. thaliana (AP31, AP36, AP37), and human (H45, H48, H49) Protox genes showed 43~65, 41~72 and 17~70-fold more resistance to oxyfluorfen, respectively, than the wild type. Among transgenic rice lines overexpressing Protox genes, several lines showed normal growth compared with the wild type, but several lines showed in reduction of plant height and shoot fresh weight under different light conditions. However, reduction of plant height of AP37 was much higher than other lines for the experimental period. On the other hand, the reduction of plant height and shoot fresh weight in the transgenic rice was higher in high light condition than in low light condition. Enhanced levels of Proto IX were observed in transgenic lines AP31, AP37, and H48 at 7 days after seeding (DAS) and transgenic lines PX, AP37, and H48 at 14 DAS relative to wild type. There were no differences in Mg-Proto IX of transgenic lines except for H41 and H48 and Mg-Proto IX monomethyl ester of transgenic lines except for MX, MX1, and PX. Although accumulation of tetrapyrrole intermediates was observed in transgenic lines, their tetrapyrrole accumulation levels were not enough to inhibit growth of transgenic rice. There were no differences in reactive oxygen species, MDA, ALA synthesizing capacity, and chlorophyll between transgenic lines and wild type indicating that accumulated tetrapyrrole intermediate were apparently not high enough to inhibit growth of transgenic rice. Therefore, the growth reduction in certain transgenic lines may not be caused by a single factor such as Proto IX, but by interaction of many other factors.

The Pulation Structure of the Pacific Cod (Gadus macrocephalus Tilesius) Based on Mitochondrial DNA Sequences (미토콘드리아 유전자 염기서열 분석에 의한 대구 계군 분석)

  • Seo, Young-Il;Kim, Joo-Il;Oh, Taeg-Yun;Lee, Sun-Kil;Park, Jong-Hwa;Kim, Hee-Yong;Cho, Eun-Seob
    • Journal of Life Science
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    • v.20 no.3
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    • pp.336-344
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    • 2010
  • To assess population structure and genetic diversity among the Pacific cod (Gadus macrocephalus), we investigated mtDNA COI gene sequences of 7 populations. Samples were obtained from Sokcho, Wolsung, Geojedo, Yeosu, Geomundo and Westsouth in 2008 and 2009 (n=28). The sequence analysis of 28 individual samples showed 8 haplotypes, ranging in sequence divergence by pairwise comparisons from 0.2 to 2.2% (1 bp-11 bp). The Gal haplotype was found in Wolsung, Geojedo, Yeosu, Geomundo and Westsouth, and was regarded as the main haplotype of Korean Pacific cod. Ga2, Ga3, Ga6 and Ga7 haplotypes were found only in Sokcho. In the PHYLIP analysis, 8 haplotypes formed two independent groups: cladeA consisted of Ga2, Ga3, Ga6 and Ga7 haplotypes, whereas cladeB contained Gal, Ga4, Ga5 and Ga8 haplotypes. The genetic relationship between the two groups was weakly supported by bootstrap analysis(<50%). In pairwise comparisons between 6 populations other than that from Sokcho, a very high per generation migration ratio ($N_m$=infinite) and a very low level of geographic distance ($F_{sr}=-0.0123-(-0.0423)$) were observed. The estimates of genetic distance between Sokcho and the other localities were all statistically significant (p<0.05, p<0.01, p<0.001), indicating a limited mtDNA-based gene flow between Sokcho and other regions. The finding of the lowest genetic diversity in the Sokcho population (nucleotide diversity=0.00589) may be a result of relatively small population size and interrupted gene flow to other localities. Consequently, the overall considerable migration of Pacific cod population in Korea caused a genetically homogeneous structure to form, although a distinct population was found in this study.

Dose Characteristics of Total-Skin Electron-Beam Irradiation with Six-Dual Electron Fields (Six-Dual 전자선 조사면에 의한 전신 피부 조사의 선량 특성)

  • Choi, Tae-Jin;Kim, Jin-Hee;Kim, Ok-Bae
    • Radiation Oncology Journal
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    • v.16 no.3
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    • pp.337-345
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    • 1998
  • Purpose : To obtain the uniform dose at limited depth to entire surface of the body, the dose characteristics of degraded electron beam of the large target-skin distance and the dose distribution of the six-dual electron fields were investigated Materials and Method : The experimental dose distributions included the depth dose curve, spatial dose and attenuated electron beam were determined with 300 cm of target-skin distance (TSD) and full collimator size (35*35 $cm^2$ on TSD 100 cm) in 4 MeV electron beam energy. Actual collimated field size of 105 cm * 105 cm at the distance of 300 cm could include entire hemibody. A patient was standing on step board with hands up and holding the pole to stabilize his/her positions for the six-dual fields technique. As a scatter-degrader, 0.5 cm of acrylic plate was inserted at 20 cm from the body surface on the electron beam path to induce ray scattering and to increase the skin dose. Results : The full width at half maximum(FWHM) of dose profile was 130 cm in large field of 105*105 $cm^2$ The width of $100\pm10\%$ of the resultant dose from two adjacent fields which were separated at 25 cm from field edge for obtaining the dose unifomity was extended to 186 cm. The depth of maximum dose lies at 5 mm and the 80$\%$ depth dose lies between 7 and 8 mm for the degraded electron beam by using the 0.5 cm thickness of acrylic absorber. Total skin electron beam irradiation (TSEBI) was carried out using the six dual fields has been developed at Stanford University. The dose distribution in TSEBI showed relatively uniform around the flat region of skin except the protruding and deeply curvatured portion of the body, which showed excess of dose at the former and less dose at the latter. Conclusion : The percent depth dose, profile curves and superimposed dose distribution were investigated using the degraded electron beam through the beam absorber. The dose distribution obtained by experiments of TSEBI showed within$\pm10\%$ difference except the protruding area of skin which needs a shield and deeply curvatured region of skin which needs boosting dose.

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Fine needle aspiration cytology of enlarged lymph nodes in children and adolescents (소아 및 청소년에 있어서 림프절 종대에 대한 미세침 흡인 세포검사법)

  • Lee, Seung Min;Oh, Yoon Jung;Jun, Yong Hoon;Hong, Young Jin;Son, Byong Kwan;Kim, Soon Ki;Han, Jee Young;Chu, Young Chae
    • Clinical and Experimental Pediatrics
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    • v.49 no.2
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    • pp.167-172
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    • 2006
  • Purpose : Palpable lymph nodes are common in the pediatric population, and most of them are either inflammatory or congenital. As the diagnostic evaluation, fine needle aspiration cytology (FNAC) is known to be more convenient and economical than tissue biopsy. We evaluated the usefulness of FNAC in children and adolescents. Methods : Four hundred and thirteen FNAC samples(M : F=1 : 1.15) were retrospectively analyzed in patients who were brought to Inha University Hospital, from August 1999 to August 2004. Results : The most common age group was 16-20 years of age(N=148, 35.8 percent). The cervical area was the most frequently involved site(N=310, 75 percent). Non-specific lymphadenitis was the most common(N=227, 54.9 percent), followed by the benign neoplasm(N=59, 14.2 percent). Malignant tumors were 18 cases(4.3 percent), and congenital diseases were found in 10 cases(2.4 percent). In inflammatory lesions, tuberculous lymphadenitis(N=22, 5.3 percent) was the most common with a histologic sensitivity of 90.9 percent. The peripheral blood and serologic studies were non-specific. Fifty nine percent(N=244) of lymphadenitis improved without specific management. Antibiotics were prescribed in 15.2 percent of lymphadenitis and lymphadenectomy was performed in 12.6 percent. Conclusion : Most of the enlarged lymph nodes in children and adolescents were benign. These results show FNAC is a safe, rapid and reliable diagnostic procedure for the appropriate differential diagnosis of enlarged lymph nodes in children and adolescents.

Pollution of Heavy Metals and Sedimentation Rate in the Sediments of Suyeong Bay, Pusan (수영만 퇴적물의 퇴적속도와 중금속 오염)

  • YANG Han-Soeb;KIM Seong-Soo
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.27 no.5
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    • pp.643-658
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    • 1994
  • The sedimentary records of anthropogenic metal loads in the Suyeong Bay, Pusan were determined by combining the Pb-210 dating technique with the measurements of heavy metals in the sediment cores. The sedimentation rates of sediment particles ranged from $0.12\;to\;0.20\;g/m^2/yr\;or\;2.4{\sim}4.0\;mm/yr$ in accumulation rates. The lowest sedimentation rate was observed at station S3 which was characterized by a bottom with relatively low organic matter contents(e.g. TIL and TOC). Heavy metals showed generally higher concentrations at station S1 and S2 near the mouth of the Suyeong River than at station S3 and the outmost station S4. The contents of copper, lead and zinc in the sediment cores especially from station S1 and S2 began to increase around 1930, and were at their highest levels in the $1960{\sim}1970$ period as a result of increasing industrial activities. Concentrations of these heavy metals have slightly decreased since 1970, probably due to regulation of pollution discharge. The natural background levels of copper, lead and zinc in the sediments of this bay ranged $18{\pm}4ppm,\;28{\pm}6ppm\;and\;74{\pm}9ppm$, respectively, by averaging the contents in the sediment depths corresponding to periods between about 1900 and 1920 at the four stations. The total amounts of anthropogenic loads deposited in the sediments since about 1930 were estimated to be $9{\sim}291{mu}g/cm^2$ for lead, $165{\sim}1122{mu}g/cm^2$ for zinc and $20{\sim}208{mu}g/cm^2$ for copper. These values were remarkably high at stations S1 and S2 relative to the other two stations. At stations S1 and S2, the anthropogenic loads of lead, copper and zinc constituted $29{\sim}30\%,\;32{\sim}42\%\;and\;28{\sim}35\%$ of the total sedimentary inventories at the present day, respectively. These metal contents have a good correlation(r>0.7) with each other and cadmium measurements also show a positive linear relation with nickel or total organic nitrogen.

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Evaluation of Sustainability for Olive Flounder Production by the Systems Ecology I. EMERGY Analysis of Olive Flounder Production (시스템 생태학적 접근법에 의한 넙치생산의 지속성 평가 I. 넙치생산에 대한 EMERGY 분석)

  • KIM Nam Kook;SON Ji Ho;KIM Jin Lee;LEE Suk Mo
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.34 no.3
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    • pp.218-224
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    • 2001
  • Olive flounder is one of the most important aquaculture species in Korea. Interest in the aquaculture of olive flounder has increased recently because of its good growth characteristics and high market price, However, the productivity of olive flounder aquaculture depends on economic inputs such as fuels, facilities, and labor, In this study, EMERGY concepts was used to compare the environment and economy of two olive flounder production methods, fishing fisheries and aquaculture, and to evaluate the sustainability of olive flounder production, EMERGY spelled with an 'm' is a universal measure of real wealth of the work of nature and society made on a common basis. Calculations of EMERGY production and storage provide a basis for making choices about environment and economy following. the general public policy to maximize real wealth, production and use. EMERGY flows from environment were $94.13\%$ for olive flounder fishing fisheries, and $2.20\%$ for aquaculture. EMERGY yield ratio, environmental loading ratio and sustainability index were 17.05, 1.02 and 274 for fishing fisheries and 0.06, 44.41 and 0.023 for aquaculture, respectively. These ratios indicate that the fishing fisheries will yield more net EMERGY, while the aquaculture requires a lower investment of EMERGY.

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