• Title/Summary/Keyword: Fragment method

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Analysis of Upstream Regulatory Region from Populus nigra × P. maximowiczii by Inverse PCR Technique (Inverse PCR 기법(技法)을 이용(利用)한 양황철 DNA의 Regulatory Region의 탐색(探索))

  • Son, Suk Gyu;Hyun, Jung Oh
    • Journal of Korean Society of Forest Science
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    • v.87 no.3
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    • pp.334-340
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    • 1998
  • This research was conducted to identify plant regulatory regions by gene tagging method. A promoterless GUS coding sequence was introduced to Populus nigra ${\times}$ P. maximowiczii via Agrobacterium strains(LBA4404/EHA101), and putative transgenic poplars were selected by culturing on medium containing G418($60mg/{\ell}$) and by GUS assay. Among them one positive plant was to amplify the native sequences flanking to the introduced GUS gene in plant genome by inverse PCR method and from this 730 by DNA product was obtained. After subcloning and sequencing, it has 88% homology to the Eucalyptus gunnii CAD(cinnamyl alcohol dehydrogenase) gene. The GUS gene fused with the putative promoter reinserted into poplar leaves by particle bombardment method to test the funtional promoter activity. Upon staining with X-gluc, many blue spots appeared on the leaf segments bombarded by the chimeric gene 2-3 days, thus the isolated DNA fragment contain some possible coding region as well as a putative regulatory sequences of poplar CAD gene.

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Development of a Monitoring System for Water-borne Bacteria by a Molecular Technique, PCR-RFLP-sequence Analysis

  • Lee, Ji-Young;Jeong, Eun-Young;Lee, Kyu-sang;Seul-Ju;Kim, Jong-Bae;Kang, Joon-Wun;Lee, Hye-Young
    • Biomedical Science Letters
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    • v.9 no.3
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    • pp.139-144
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    • 2003
  • Since water borne infection causes acute diseases and results in spread of diseases by secondary infection, the prevention is very important. Therefore, it is necessary to have a method that is rapid and effective to monitor pathogenic bacteria in drinking water. In this study, we employed a systematic method, Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) analysis, to develop an effective monitoring system for possible bacterial contaminants in drinking water. For this purpose, PCR primers were derived from 992 bp region of the 16s rRNA gene that is highly conserved through the different species of prokaryotes. To test whether the PCR primers designed are indeed useful for detecting all the possible microbial contaminants in the water, the primers were used to amplify 16s rRNA regions of different microbial water-borne pathogens such as E. coli, Salmonella, Yersinia, Listeria, and Staphylococcus. As expected, all of tested microorganisms amplified expected size of PCR products indicating designed PCR primers for 16s rRNA indeed can be useful to amplify all different microbial water-borne pathogens in the water. Furthermore, to test whether these 16s rRNA based PCR primers can detect bacterial populations present in the water, water samples taken from diverse sources, such as river, tap, and sewage, were used for amplification. PCR products were for then subjected for cloning into a T-vector to generate a library containing 16s rRNA sequences from various bacteria. With cloned PCR products, RFLP analysis was done using PCR products digested with restriction enzyme such as Hae III to obtain species-specific RFLP profiles. After PCR-RFLP, the bacterial clones which showed the same RFLP profiles were regarded as the same ones, and the clones which showed distinctive RFLP profiles were subsequently subjected for sequence analysis for species identification. By this PCR-RFLP analysis, we were able to reveal diverse populations of bacteria living in water. In brief, in unsterilized natural river water, over 60 different species of bacteria were found. On the other hand, no PCR products were detected in drinking tap-water. The results from this study clearly indicate that the PCR-RFLP-sequence analysis can be a useful method for monitoring diverse, perhaps pathogenic bacteria contaminated in water in a rapid fashion.

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Effect of Different Milling Methods on Distribution of Particle Size of Rice Flours (제분방법이 쌀가루의 입자크기에 미치는 영향)

  • Kum, Jun-Seok;Lee, Sang-Hyo;Lee, Hyun-Yu;Kim, Kil-Hwan;Kim, Young-In
    • Korean Journal of Food Science and Technology
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    • v.25 no.5
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    • pp.541-545
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    • 1993
  • Two different methods (Sieve shaker, Elzone particle size analyzer) were used to investigate rice flour particle size obtained by various milling method. Results of Elzone particle size analyzer were more effective than Sieve shaker in determining particle size, and the distribution of particle size of rice flours was affected by the type of the milling methods used. A rice flour, prepared in a Pin mill had a particle size range of $60{\sim}500$ mesh, and 30.38% of the sample was in the particle size range $200{\sim}270$ mesh. A rice flour, prepared in a Colloid mill had a particle size range of $40{\sim}500$ mesh and more of flour particles appeared in the range $140{\sim}200$ mesh than any other particle size. A rice flour, prepared in a Micro mill had a particle size range of $140{\sim}500$ mesh, and 41.62% of the sample was in the particle size range over 500 mesh. A rife flour, prepared in a Jet mill had a finer flour particle size was over the particle size range 500 mesh. The finer rice flour gave the highest L value and the lowest a value. The wet-milled flour particles were observed as a cluster of starch granules and the particles of rice flour (dry-milling) were observed as fragment of rice grains. Scanning Electron Photomicrographs revealed that visual differences in structure between milling methods, and similar results with Elzone particle size analyzer method in particle size.

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Anterior Instability of the Knee after Arthroscopic Treatment for Tibial Spine Fractures (경골극 골절의 관절경적 치료 후 슬관절의 전방 불안정성)

  • Choi, Chong-Hyuk;Kim, Sung-Hwan;Joo, Jong-Hwan;Kim, Hyung-Sik
    • Journal of the Korean Arthroscopy Society
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    • v.10 no.1
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    • pp.26-32
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    • 2006
  • Purpose: We evaluated the result of the degree of reduction and anterior instability after arthroscopic treatment of tibial spine fractures. Materials and Methods: Thirty-two cases of tibial spine fractures treated with arthroscopic procedure could be evaluated and the mean follow-up period was $18\;(12{\sim}48)$ months. Evaluations included Lachman test, rotatory instability examination and side to side difference (SSD) using KT-2000 arthrometer. The degree of radiologic reduction was classified as accurate reduction, anterior elevation and reduction failure, according to which the degree of anterior instability was measured. Results: There were twenty three cases (71%) of accurate duction, eight cases (25%) of anterior elevation and one case (4%) of education failure. Eight (29%) of twenty eight cases with pull-out method showed anterior elevation. The SSDs of the accurate reduction and the anterior elevation group were 2.0 mm and 2.8 mm, respectively showing no significant difference. However, the percent age of grade 0 Lachman test and SSD less than 3 mm of the accurate reduction group were significantly higher. Conclusion: Accurate reduction is important in reducing anterior instability especially in unstable fractures. Therefore, arthroscopic pull-out method should be performed with great caution to avoid anterior elevation of the fracture fragment.

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Distribution of Wildbirds According to Habitat Environment in Gap Stream (갑천의 서식지 환경에 따른 야생조류 분포에 관한 연구)

  • Lee, Joon-Woo;Lee, Do-Han;Paik, In-Hwan
    • Korean Journal of Agricultural Science
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    • v.30 no.1
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    • pp.41-58
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    • 2003
  • This study was conducted to investigate bird community and to suggest a proper way how to manage protect bird community in Gap stream. The survey was carried out over four sections by the line transect method and point counts method from September 2001 to August 2002. Natural stream region as Gasuwon Bridge - Mannyeon Bridge are observed birds were 11 orders 29 families 67 species, Artificial stream region as Mannyeon Bridge - Daedeok Bridge are observed birds were 6 orders 10 families 30 species, Daedeok Bridge - Wonchon Bridge are 8 orders 12 families 28 species, Wonchon Bridge - Gap Stream Bridge are 8 orders 18 families 40 species. All the observed birds in artificial stream region are 8 orders 19 families 47 species. Number of species in natural stream region was higher than artificial stream region owe to a various habitat environment such as forest, cultivated land, streamside forest, sandy plain, gravelly field, reedy field etc. and can not add with the interface and the usage of the human. Number of species in artificial stream region was lower than natural stream region owe to a simple habitat environment and the water ecosystem is severed with embankment block and grass plot with the land ecosystem. The furtherance of various habitat environment which considers the ecosystem like the natural stream as the water ecosystem is joined together with the land ecosystem is desired to attract various wildbirds in Gap stream. The design is desired with the maintenance of the stream to consider the stream corridor which plays ecological important role as connect the fragment habitats.

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Tetrodotoxin in a Pufferfish, Fugu xanthopterus (Korean Name, Ggachibog) (까치복(Fugu xanthopterus)의 독성)

  • Hyun-Dae Kim;Yeung-Ho Park;Dong-Soo Kim
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.23 no.3
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    • pp.502-508
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    • 1994
  • A total of 24 specimens of the pufferfish, Fugu xanthopterus, purchased at a fixhmarket in Pusan, korea were examined for toxicity using the assay method of tetrodotoxin (TTX). Also, the toxins isolated from the puffer liver were partially purified and analyzed for their chemical composition by instrumental behaviors. On the whole, when the level of toxicity in each organ was analyzed compared to that of liver, they were 100 % of the lover, 92 % for the intestine, 75% for the skin, 17% for the muscle, 785 for the testis, 87% for the ovary, and 71% for bile. The highest and average scores of toxicity for the liver were 917 and $231{\pm}51MU/g$ liver, respectively. The toxins of the puffer gave four peaks in HPLC whose retention times (10, 20, 22 and 25 min) were close to those of TDA, TTX, 4-epi-TTX, and and -TTX, respectively.

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Production of Transgenic Goats by Sperm-mediated Exogenous DNA Transfer Method

  • Zhao, Yongju;Wei, Hong;Wang, Yong;Wang, Lingbin;Yu, Mingju;Fan, Jingsheng;Zheng, Shuangyan;Zhao, Cong
    • Asian-Australasian Journal of Animal Sciences
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    • v.23 no.1
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    • pp.33-40
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    • 2010
  • In this study, the production of transgenic goats using sperm to integrate exogenous DNA and artificial insemination (AI) was carried out and the technical protocols for sperm-mediated gene transfer (SMGT) in the goat were optimized. The standard sperm parameters and the ability to bind foreign genes were assessed to select suitable sperm donor bucks. A total of 134 oestrous does were divided into 4 groups and inseminated using different methods and sperm numbers. The does of Groups I to III were inseminated with fresh semen ($1-2\times10^{7}$ and $10^{6}$ sperm) or frozen-thawed semen ($10^{6}$ sperm), respectively, through conventional intra-cervical AI, and the does of Group IV with frozen-thawed semen ($10^{6}$ sperm) through intrauterine AI. Total genomic DNAs were extracted from ear biopsies of the offspring. The presence of $pEGFP-N_{1}$ DNA was screened by PCR and then by Southern blotting analysis. A total of 76 live kids were produced and 8 kids were tested transgene positive on the basis of agarose gel electrophoresis of the PCR-amplified fragment. Southern blotting analysis of the samples showed 5 positive kids. A transgenic ratio of 10.53% was detected using PCR and 6.58% using Southern blotting. The positive kid rate assayed by PCR and Southern blotting of frozen-thawed goat semen was 3.61% and 9.27% higher than that of untreated semen. The results show that transgenic goats can be produced efficiently by the method of artificial insemination using sperm cells to integrate the exogenous DNA and intrauterine insemination allowed low numbers of DNA-transfected spermatozoa to be used, with satisfactory fertility.

Rapid Detection of the pathogenic agent of Bacterial white enteritis of Larval and Juvenile Stages in Olive flounder (Paralichthys olivaceus) (넙치(Paralichthys olivaceus)자치어 장관백탁증(Bacterial white enteritis) 원인균의 신속 검출)

  • Mun, Yeong-Geon;Park, Geun-Tae;Son, Hong-Ju;Lee, Sang-Hyeon;Lee, Jeong-Min;Heo, Mun-Su
    • Journal of fish pathology
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    • v.17 no.3
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    • pp.159-169
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    • 2004
  • Bacterial wihte enteritis ocurred by infection of V. ichthyoenteri is a devastating disease in olive flounder (Paralichthys olivaceus) hatcheries in Korea. Since white enteritis has been a problem in aquqtic industries, necessity of a rapid detection method is increased. In an attempt to develop rapid PCR method the detection of V. ichthyoenteri, we examined the 16S-23S rRNA intergenic spacer region(ISR) of V. ichthyoenteri and developed species-specific primer for V. ichthyoenteri. The intergenic spacers were amplified by primers complementary to conserved region of 16S and 23S rRNA genes. The intergenic spacer region between the 16S and 23S rRNA genes of V. ichthoenteri were investigated by PCR fragment length typing and DNA sequencing. Analysis of the ISR sequences showed that V. ichthyoenteri contains one types of polymorphic ISRs. The size of ISRs ranged 348bp length and not contains tRNA genes. Mutiple alignment of representative sequences from different Vibrio species revealed several domains of high sequence variability, and allowed to design species-specific primer for detection of Vibrio ichthyoenteri. PCR. The specific of the primer was examined using genomic DNA prepared from 19 different Vibrio species, isolated 18group Vibrio species. The results showed that the PCR reaction using species-specific primer designed in this study can be used to detect V. ichthyoenteri.

gyrA Mutations Found Among Ofloxacin-resistant Mycobacterium tuberculosis is Isolated from Korea

  • Kim Junho;Kim Yeun;Bae Kiho;Song Taek-Sun;Cho Sang-Nae;Lee Hyeyoung
    • Biomedical Science Letters
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    • v.11 no.4
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    • pp.465-471
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    • 2005
  • Ofloxacin has antimycobacterial activity that possibly contributes a pivotal role in the second-line drug regimens that are used for the treatment of multidrug-resistant tuberculosis. However, in some communities, the resistance rate of Mycobacterium tuberculosis to this agent is surging. Therefore, a rapid and accurate method that can be used to determine the resistance of M tuberculosis to the ofloxacin can be very useful for effective treatment of the patients. As an effort to develop such a method, this study was set up to reveal general types of mutations that are related to ofloxacin resistance of M tuberculosis. From previous studies, it has been well known that ofloxacin resistance is associated with mutations in a gene encoding the gyrase A subunit protein. In this study, we obtained 43 ofloxacin-resistant and 50 ofloxacin-susceptible M tuberculosis clinical isolates from Masan National TB Hospital, and sequences of DNA fragment of 320 bp, region of gyrA corresponding to the ofloxacin resistance-determining region were analyzed. In brief, the results showed that a total of seven mutation types were found at gyrA. Theses mutations were all clustered within nucleotides 2574 to 2586 of the gyrA gene (codons 88 to 94). Codon 94 was the most frequently substituted site. Twenty-four of the 43 isolates had mutations at this position resulting in a total of five different types of amino acid changes $(Asp{\to}Ala,\;Asp{\to}Gly,\;Asp{\to}His,\;Asp{\to}Tyr,\;and\;Asp{\to}Asn)$. Five isolates contained a mutation at codon 90 resulting $Ala{\to}Val$ change. Four isolates had mutations at codon 91 causing a $Ser{\to}Pro$ change at this site. Two isolates contained a mutation at codon 88 and each of them resulted in different types of amino acid changes $(Gly{\to}Cys,\;Gly{\to}Ala)$. On the other hand, polymorphic site at codon 95 was found in both ofloxacin-resistant and ofloxacin-susceptible isolates. From these results, we concluded that the rate of mutations present in gyrA among ofloxacin-resistant M. tuberculosis in Korea is similar to the general rates of mutations found throughout the world. Subsequently, an oligonucleotide probe was designed based on the results of sequence analysis and was used to develop a dot blot hybridization assay system to determine ofloxacin-resistance of M tuberculosis. To evaluate this probe, dot-blot hybridization was carried out using other 57 clinical isolates, and the results showed that the dot-blot hybridization assay is good for detecting sequence alterations atgyrA gene.

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A Comparative Study of the Expantionist Architectural Concept in Post-Modern Museums -focused on Neue Staatsgalerie Stuttgart and Stadtisches Museum Abteiberg Monchengladback in Germany- (포스트모던 미술관 건축의 확장된 건축개념에 관한비교연구 -독일 스튜트가르트 국립미술관 신관과 묀헨글라드바하 시립미술관을 중심으로-)

  • 김명옥
    • Korean Institute of Interior Design Journal
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    • no.14
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    • pp.82-89
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    • 1998
  • During the boom of museum building in the 1970s-80s two museum wee erected in Germany. They are James Stirling's Neue Stattsgalerie Stuttgart and hans Hollein's Stadtisches Museum Abteiberg Monchengladbach, These two museums share the folowing point of similarity : Stirling's Neue Stattsgalerie Stuttgart and Hollein's Staditisches Museum Abetiberg Monchenbladbach both manifest the conditions of the times in light of their respective cities' local characters and historical contexts without being bound to traditional formality. Stirling and Hollein attempted to grasp the meaning of the 기nuseum as a city in miniature." Taking into account the t two museums’s territorial characteristics of being situated on a slope, both Stirling and Hollein made the walker to pass t through their building complex and introduced the concept of a public square within them. As a result, the museums are not j just two large buildings but are architecture composed of a collage of various structures. S Stirling’s architecture employs the method of attaching additional elements on top of basic constituents, which is suggestive '||'&'||'#61551; of the historical fragment from Shinkel's Altes Museum. On the other hand, Hollein applies a collage-style method as if he w were doing urban planning, maintaining the distinctiveness of each of the various forms and materials of buildings. T The object style buildings of the two museums actively demonstrate the contrast of double meaning to represent the a ambiguous and multifarious characteristics of the modern times. Stirling explores the theme of opposition and coexistence in h his Neue Stattsgalerie Stuttgart by placing a series of opposing concepts, such as the past and present and histor${\gamma}$ and t technology, in one space. Thereby, the contradiction and its appeal are manifested. Hollein made use of the visual illusion and c contradiction in alluding to the irony of the modern reality induced by nature and culture and history and technology. F For the above reasons and methods, James Stirling ’ s Neue Stattsgalerie Stuttgart allowed the penetration of art into daily l life and became the general public’s favored museum for its free and unrestrained environment. Likewise, Hans Hollein’s S Stadtisches Museum Abteiberg Moncbengladbach contribute to the development of museum function as it an expression of art i in the form of a complex piece of scupture on its own.ts own.

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