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Integrated Rotary Genetic Analysis Microsystem for Influenza A Virus Detection

  • Jung, Jae Hwan;Park, Byung Hyun;Choi, Seok Jin;Seo, Tae Seok
    • Proceedings of the Korean Vacuum Society Conference
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    • 2013.08a
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    • pp.88-89
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    • 2013
  • A variety of influenza A viruses from animal hosts are continuously prevalent throughout the world which cause human epidemics resulting millions of human infections and enormous industrial and economic damages. Thus, early diagnosis of such pathogen is of paramount importance for biomedical examination and public healthcare screening. To approach this issue, here we propose a fully integrated Rotary genetic analysis system, called Rotary Genetic Analyzer, for on-site detection of influenza A viruses with high speed. The Rotary Genetic Analyzer is made up of four parts including a disposable microchip, a servo motor for precise and high rate spinning of the chip, thermal blocks for temperature control, and a miniaturized optical fluorescence detector as shown Fig. 1. A thermal block made from duralumin is integrated with a film heater at the bottom and a resistance temperature detector (RTD) in the middle. For the efficient performance of RT-PCR, three thermal blocks are placed on the Rotary stage and the temperature of each block is corresponded to the thermal cycling, namely $95^{\circ}C$ (denature), $58^{\circ}C$ (annealing), and $72^{\circ}C$ (extension). Rotary RT-PCR was performed to amplify the target gene which was monitored by an optical fluorescent detector above the extension block. A disposable microdevice (10 cm diameter) consists of a solid-phase extraction based sample pretreatment unit, bead chamber, and 4 ${\mu}L$ of the PCR chamber as shown Fig. 2. The microchip is fabricated using a patterned polycarbonate (PC) sheet with 1 mm thickness and a PC film with 130 ${\mu}m$ thickness, which layers are thermally bonded at $138^{\circ}C$ using acetone vapour. Silicatreated microglass beads with 150~212 ${\mu}L$ diameter are introduced into the sample pretreatment chambers and held in place by weir structure for construction of solid-phase extraction system. Fig. 3 shows strobed images of sequential loading of three samples. Three samples were loaded into the reservoir simultaneously (Fig. 3A), then the influenza A H3N2 viral RNA sample was loaded at 5000 RPM for 10 sec (Fig. 3B). Washing buffer was followed at 5000 RPM for 5 min (Fig. 3C), and angular frequency was decreased to 100 RPM for siphon priming of PCR cocktail to the channel as shown in Figure 3D. Finally the PCR cocktail was loaded to the bead chamber at 2000 RPM for 10 sec, and then RPM was increased up to 5000 RPM for 1 min to obtain the as much as PCR cocktail containing the RNA template (Fig. 3E). In this system, the wastes from RNA samples and washing buffer were transported to the waste chamber, which is fully filled to the chamber with precise optimization. Then, the PCR cocktail was able to transport to the PCR chamber. Fig. 3F shows the final image of the sample pretreatment. PCR cocktail containing RNA template is successfully isolated from waste. To detect the influenza A H3N2 virus, the purified RNA with PCR cocktail in the PCR chamber was amplified by using performed the RNA capture on the proposed microdevice. The fluorescence images were described in Figure 4A at the 0, 40 cycles. The fluorescence signal (40 cycle) was drastically increased confirming the influenza A H3N2 virus. The real-time profiles were successfully obtained using the optical fluorescence detector as shown in Figure 4B. The Rotary PCR and off-chip PCR were compared with same amount of influenza A H3N2 virus. The Ct value of Rotary PCR was smaller than the off-chip PCR without contamination. The whole process of the sample pretreatment and RT-PCR could be accomplished in 30 min on the fully integrated Rotary Genetic Analyzer system. We have demonstrated a fully integrated and portable Rotary Genetic Analyzer for detection of the gene expression of influenza A virus, which has 'Sample-in-answer-out' capability including sample pretreatment, rotary amplification, and optical detection. Target gene amplification was real-time monitored using the integrated Rotary Genetic Analyzer system.

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Immunogenicity of Synthetic Peptide Specific for Major Immunogenic Determinat of Hepatitis B Surface Antigen (B형간염(型肝炎) 표면항원(表面抗原)의 주면역원(主免疫原) 결정기(決定基)에 특이(特異)한 합성(合成) Peptide의 면역원성(免疫原性)에 관한 연구(硏究))

  • Shin, Kwang-soon;Han, Su-nam
    • Korean Journal of Veterinary Research
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    • v.25 no.1
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    • pp.7-17
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    • 1985
  • Many investigators have been pursuing various attempts so far to produce hepatitis B surface antigen(HBsAg) vaccines using the techniques such as isolation from plasma of chronic HBsAg carrier, recombinant DNA technique or preparation of synthetic peptides specific for immunogenic determinants. Hepatitis B virus can not grow on any cell lines by the tissue culture technique at the present time. The plasma of chronic HBsAg carrier is expensive and its source is limited. The HBsAg from the recombinant DNA technique gave still very low yield. Another approach, therefore, has been initiated to develop a synthetic hepatitis B virus vaccine. The possible use of several distinct synthetic vaccines in prophylaxis can be facilitated by availability of full synthetic immunogens. Peptides synthesized for potential application as antiviral vaccines have been mostly tested in the form of conjugates with carrier proteins, although the free synthetic peptide can be immunogenic. To understand basic knowledges on the antigenicity and immunogenicity of a synthetic peptide specific for major immunogenic determinant of HBsAg, a nonapeptide, $H_2N^{139}Cys-Thr-Lys-Pro-Thr-Asp-Gly-^{146}Asn-Aba$ COOH, which corresponds to HBsAg amino acid residues 139 to 147, was synthesized by the Merrifield's solid-phase method with a slight modification. The antigenicity and immunogenicity of this specific synthetic peptide were examined comparing with purified plasma-derived natural HBsAg. The results obtained are as follows; 1. The peptide synthesized showed the identical amino acid composition to the theoretical value. The degree of purification and molecular weight were acertained by methods of high performance liquid chromatography and mass spectrometry. 2. Using m-maleimidobenzoyl-N-hydroxysuccinimide ester as a conjugating agent, the synthetic peptide was conjugated to rabbit albumin and ${\gamma}$-globulin, tetanus and diphtheria toxoids, and keyhole limpet hemocyanin. Their conjugation yields were 8.3, 9.5, 15.8, 13.5, and 11.2%, respectively. 3. The natural HBsAg was purified from plasma of chronic HBsAg carrier. By the electron microscopic observation of the purified natural HBsAg preparation, no Dane particles were observed and the preparation showed negative DNA polymerase activity. 4. Antigenicity of the synthetic peptide and the plasma-derived natural HBsAg was determined by competition radioimmunoassay using $^{125}I$-natural HBsAg. Their 50% inhibitions appeared as $90{\mu}g/ml$ and $0.12{\mu}g/ml$ for the synthetic peptide and the natural HBsAg, respectively. This indicates that the former was about 750-fold less antigenic than the latter. 5. Immunogenicity of the synthetic peptide was determined by administering the peptide-carrier conjugates into rabbits with and without Freund's complete adjuvant. Regardless the carrier proteins and adjuvant, positive immune responses to the synthetic peptide were observed. The higher antibody titers, however, were shown in the groups administered with Freund's complete adjuvant. 6. Immunizing dose 50% in mice of the various peptide-carrier conjugates was 5.47, 6.00, 65.16, 31.25 and $13.03{\mu}g/dose$ for rabbit albumin and ${\gamma}$-globulin, tetanus and diphtheria toxoids, and keyhole limpet hemocyanin, respectively, while the natural HBsAg showed $0.65{\mu}g/dose$. 7. It was postulated that homologous proteins prefer to heterologous ones as the carriers.

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The Seosan County Family Planning/Maternal & Child Health Service Research Project, Korea -Project Design and Findings of the Baseline Survey- (가족계획(家族計劃) 및 모자보건사업(母子保健事業)의 효율적 통합방안(統合方案)에 관한 연구(硏究)(서산군(瑞山郡)) -기초조사보고(基礎調査報告)-)

  • Bang, S.;Cho, T.H.;Lee, S.J.;Han, S.H.;Lim, K.J.;Ahn, M.Y.
    • Journal of Preventive Medicine and Public Health
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    • v.16 no.1
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    • pp.163-192
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    • 1983
  • In order to facilitate the Korean government's efforts in integrating family planning and maternal & child health at the primary health care level (or township level), the Soon Chun Hyang College of Medicine, with the financial and technical assistance of WHO, has under-taken a service research project. The project has employed a quasi-experimental study design introducing interventions tat provide crucial factors lacking in the ongoing government programs such as midwives and qualified referral physicians. The study is being conducted in three locations, one control area and two study areas. Before introducing trained Nurse/Midewives into the study areas, a baseline prevalence survey was undertaken from 15 July 1981 to 10 August 1981 in selelcted townships of Seosan County. In this sample survey of bath the study and control areas, 2,484 eligible women (97% reponse rate) were interviewed to obtain benchmark data on basic evaluation indicators related to family planning and maternal and child health. The salients results were summarized as follows.: 1. CONTACT RATES WITH HEALTH WORKERS; During the year preceding the survey, 12% of women were visited by government health workers. The primary reason for such visits by health workers was family planning (45% of the visits). About 34% of the women visited the health centers during the year. The primary reason for visiting health centers was immunizations for their children (45% of the visits). 3. FAMILY PLANNING USE RATE; The baseline data showed little difference between women in the study area and the control area on contraceptive use. Approximately 59% were currently using some methods. However, among those current users, almost half were practicing less effective methods of birth control such as rhythm or withdrawal. Among other methods, the tubectomy was the most popular (16%), while use of the IUD, oral pill and condom together reached only 14%. 3. PRENATAL CARE RATE; About 75% of the women reported no prenatal care for their last births (the youngest child of each women), Additionally, among women received prenatal care, over half had only one visit. 4. ATTENDANCE AT DELIVERY; Most of the women surveyed (over 80%) were attended by a non-medical person during their last delivery. These figures are somewhat comparable to the national figure of 84% for remote areas. 5. POSTNATAL CARE; The proportion of women reporting postnatal care was only 4.5%, and postnatal care was not received by the majority of women surveyed. 6. CHILD HEALTH CARE: In contrast to the low rate of maternity care for women themselves, most women reported obtaining immunization care for their children. About 75% of the women obtained Polio and/or DPT, 58% BCG, and 44% Measles vaccine for their children. However, in terms of illness care, while 35% of the women stated that their youngest child had been sick during the month preceding the survey, only 28% of these women took their child to the clinic for treatment. 7. COMPLICATIONS OF PREGNANCY AND DELIVERY AND ABNORMALITIES IN THE NEWBORN; Among all last deliveries, 18% of the women had pregnancy complications and 9% of the women had complications during delivery About 5% of the women reported abnormality in their most recent newborn. 8. REPRODUCTION EFFICIENCY; PERINATAL MORTALITY AND INFANT MORTALITY Based on data from the pregnancy history in this survey, reproduction efficiency was estimated. Out of the 11,154 pregnancies reported by all women surveyed, foetal loss was 21% (almost 16% were induced abortions) and infant deaths before reaching one year old were 3.1%. The reproduction efficiency was, therefore, reduced to 76%. In terms of perinatal and infant mortality rates, the former was 40.2 per 1,000 total births and the latter was 39.3 per 1,000 live births. Both rates described J shaped relationships with age of mothers and parity, and they were also correlated with birth interval and mother's education. In summary, this baseline survey data indicated a need for (1) improving contraceptive practices with more effective methods to prevent unwanted pregnancies and (2) providing better services for maternal and child care to protect wanted pregancies. In the Korean rural setting. the author believes that the latter is more important as the value of each child has increased as a result of the family planning campaign for the past two decades. This calls for more effective integration of Family Planning and MCH programmes to meet the needs of the family in each stage of the child bearing and rearing period with deploying more qualified personnel than the current government program personnel.

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Analysis of Respiratory Motion Artifacts in PET Imaging Using Respiratory Gated PET Combined with 4D-CT (4D-CT와 결합한 호흡게이트 PET을 이용한 PET영상의 호흡 인공산물 분석)

  • Cho, Byung-Chul;Park, Sung-Ho;Park, Hee-Chul;Bae, Hoon-Sik;Hwang, Hee-Sung;Shin, Hee-Soon
    • The Korean Journal of Nuclear Medicine
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    • v.39 no.3
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    • pp.174-181
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    • 2005
  • Purpose: Reduction of respiratory motion artifacts in PET images was studied using respiratory-gated PET (RGPET) with moving phantom. Especially a method of generating simulated helical CT images from 4D-CT datasets was developed and applied to a respiratory specific RGPET images for more accurate attenuation correction. Materials and Methods: Using a motion phantom with periodicity of 6 seconds and linear motion amplitude of 26 mm, PET/CT (Discovery ST: GEMS) scans with and without respiratory gating were obtained for one syringe and two vials with each volume of 3, 10, and 30 ml respectively. RPM (Real-Time Position Management, Varian) was used for tracking motion during PET/CT scanning. Ten datasets of RGPET and 4D-CT corresponding to every 10% phase intervals were acquired. from the positions, sizes, and uptake values of each subject on the resultant phase specific PET and CT datasets, the correlations between motion artifacts in PET and CT images and the size of motion relative to the size of subject were analyzed. Results: The center positions of three vials in RGPET and 4D-CT agree well with the actual position within the estimated error. However, volumes of subjects in non-gated PET images increase proportional to relative motion size and were overestimated as much as 250% when the motion amplitude was increased two times larger than the size of the subject. On the contrary, the corresponding maximal uptake value was reduced to about 50%. Conclusion: RGPET is demonstrated to remove respiratory motion artifacts in PET imaging, and moreover, more precise image fusion and more accurate attenuation correction is possible by combining with 4D-CT.

Effect of Addition of Lactic Acid Bacteria and Chlorella on Nutritive Values and Quality of Italian Ryegrass-Hairy Vetch Silage (젖산균과 클로렐라 첨가가 이탈리안라이그라스-헤어리베치 사일리지의 사료가치 및 품질에 미치는 영향)

  • Choi, Ki Choon;Arasu, M. Valan;Ilavenil, S.;Park, Hyung Su;Jung, Min Woong;Kime, Ji Hye;Jung, Jeong Sung;Hwangbo, Soon;Kim, Won Ho;Lim, Young Chul
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.34 no.1
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    • pp.39-44
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    • 2014
  • This study was conducted to investigate the effect of the addition of lactic acid bacteria (LAB) and chlorella on the change of nutritive value, quality and microbes regarding the mixture silage of Italian ryegrass and hairy vetch (IRG-HV silage). The content of crude protein of IRG-HV silage in only the LAB treatment was not different as compared to the control. However, the content of crude protein significantly increased in the treatment of LAB with chlorella, compared to both the control and LAB treatments (p<0.05). Acid detergent fiber, neutral detergent fiber, total digestible nutrient and in vitro dry matter digestibility of IRG-Alfalfa silage were not significantly different in all treatments. Further, the pH of IRG-HV silage in only the LAB treatment was not different as compared to control, but decreased in both LAB and chlorella treatments (p<0.05). The content of lactic acid in the LAB treatment increased as compared to the control, and also significantly increased in the LAB treatment with chlorella (p<0.05). In addition, the number of LAB in LAB treatment increased as compared to the control, and significantly also increased in the LAB treatment with chlorella (p<0.05). Therefore, we suggest that IRG-HV silage improved with addition of both LAB and chlorella.

Effect of gomchwi (Ligularia fischeri) extract against high glucose- and H2O2-induced oxidative stress in PC12 cells (PC12 신경세포에서 고당 및 과산화수소로 유도된 산화적 스트레스에 대한 곰취 추출물의 효과)

  • Park, Sang Hyun;Park, Seon Kyeong;Ha, Jeong Su;Lee, Du Sang;Kang, Jin Yong;Kim, Jong Min;Lee, Uk;Heo, Ho Jin
    • Korean Journal of Food Science and Technology
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    • v.48 no.5
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    • pp.508-514
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    • 2016
  • Effects of the ethyl acetate fraction from gomchwi (Ligularia fischeri) extract against high $glucose/H_2O_2-induced$ oxidative stress and in vitro neurodegeneration were investigated to confirm the physiological property of the extract. The ethyl acetate fraction of gomchwi extract showed the highest total phenolic contents than the other solvent fractions. An anti-hyperglycemic activity of the ethyl acetate fraction was evaluated using the ${\alpha}-glucosidase$ inhibitory assay, and the half maximal inhibitory concentration ($IC_{50}$) value for ${\alpha}-glucosidase$ was found to be $727.64{\mu}g/mL$. In addition, the ethyl acetate fraction showed excellent 2,2'-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) diammonium salt radical scavenging activity, and inhibition of malondialdehyde production. The ethyl acetate fraction also decreased intracellular reactive oxygen species, whereas neuronal cell viability against high glucose/$H_2O_2$-induced cytotoxicity was found to be increased. Finally, 3,5-dicaffeoylquinic acid as a main phenolic compound in the ethyl acetate fraction was analyzed by high-performance liquid chromatography. These results suggest that gomchwi might be a good natural source of functional materials to prevent diabetic neurodegeneration.

Effects on senior fitness, cognitive function and daily living activity of female elderly with light dementia by continuous and intermittent exercises (연속적.간헐적 운동이 경증치매 여성고령자의 기능적 체력, 인지기능 및 일상생활수행능력에 미치는 영향)

  • Kang, Joo-Seong;Jeang, Il-Hong;Yang, Jum-Hong
    • Journal of the Korea Academia-Industrial cooperation Society
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    • v.11 no.11
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    • pp.4234-4243
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    • 2010
  • Dementia such as poor concentration, anxiety and tension makes it hard to continue exercise in reality. For this, intermediate exercise is suggested in this study. And the study investigates the effects on senior fitness, cognitive function(MMSE-K) and daily living activity(ADL) by continuous exercise and intermittent exercise, which helps to provide proper exercise treatment to them. For this, female elderly patients at A dementia hospital in B Metropolitan city are selected and they have been diagnosed with possible Alzheimer's disease according to DSM-IV. Among them, six(6) are grouped for continuous exercise and five(5) for intermediate exercise, total 11 people are finally tested. They are given hand&foot exercise, Korean folk dance and band exercise three times a week for the total 12 weeks. The continuous exercise group does their exercise one time of 30 minutes a day while the intermediate exercise group for three times of each 10 minute a day. For the result, SPSS Ver. 18.0 is used to get mean value(M) and standard deviation(SD) and in order to verify the interaction effect between exercise group and time, two-way repeated ANOVA is applied and statistical significance level is set at .05. The result shows that there is significant difference in time between senior fitness and cognitive function. But there is no significant difference in group and time${\times}$group. And there is no significant difference in time, group and time${\times}$group for daily living activity. Continuous exercise group and intermediate exercise group both have the similar effects. That does not mean that intermediate exercise is the best for all people with dementia, but in terms of exercise time, intermittent exercise may be effective for patients of light dementia.

Investigation of Intertidal Zone using TerraSAR-X (TerraSAR-X를 이용한 조간대 관측)

  • Park, Jeong-Won;Lee, Yoon-Kyung;Won, Joong-Sun
    • Korean Journal of Remote Sensing
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    • v.25 no.4
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    • pp.383-389
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    • 2009
  • The main objective of the research is a feasibility study on the intertidal zone using a X-band radar satellite, TerraSAR-X. The TerraSAR-X data have been acquired in the west coast of Korea where large tidal flats, Ganghwa and Yeongjong tidal flats, are developed. Investigations include: 1) waterline and backscattering characteristics of the high resolution X-band images in tidal flats; 2) polarimetric signature of halophytes (or salt marsh plants), specifically Suaeda japonica; and 3) phase and coherence of interferometric pairs. Waterlines from TerraSAR-X data satisfy the requirement of horizontal accuracy of 60 m that corresponds to 20 cm in average height difference while current other spaceborne SAR systems could not meet the requirement. HH-polarization was the best for extraction of waterline, and its geometric position is reliable due to the short wavelength and accurate orbit control of the TerraSAR-X. A halophyte or salt marsh plant, Suaeda japonica, is an indicator of local sea level change. From X-band ground radar measurements, a dual polarization of VV/VH-pol. is anticipated to be the best for detection of the plant with about 9 dB difference at 35 degree incidence angle. However, TerraSAR-X HH/TV dual polarization was turned to be more effective for salt marsh monitoring. The HH-HV value was the maximum of about 7.9 dB at 31.6 degree incidence angle, which is fairly consistent with the results of X-band ground radar measurement. The boundary of salt marsh is effectively traceable specifically by TerraSAR-X cross-polarization data. While interferometric phase is not coherent within normal tidal flat, areas of salt marsh where the landization is preceded show coherent interferometric phases regardless of seasons or tide conditions. Although TerraSAR-X interferometry may not be effective to directly measure height or changes in tidal flat surface, TanDEM-X or other future X-band SAR tandem missions within one-day interval would be useful for mapping tidal flat topography.

Estimation of Annual Trends and Environmental Effects on the Racing Records of Jeju Horses (제주마 주파기록에 대한 연도별 추세 및 환경효과 분석)

  • Lee, Jongan;Lee, Soo Hyun;Lee, Jae-Gu;Kim, Nam-Young;Choi, Jae-Young;Shin, Sang-Min;Choi, Jung-Woo;Cho, In-Cheol;Yang, Byoung-Chul
    • Journal of Life Science
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    • v.31 no.9
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    • pp.840-848
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    • 2021
  • This study was conducted to estimate annual trends and the environmental effects in the racing records of Jeju horses. The Korean Racing Authority (KRA) collected 48,645 observations for 2,167 Jeju horses from 2002 to 2019. Racing records were preprocessed to eliminate errors that occur during the data collection. Racing times were adjusted for comparison between race distances. A stepwise Akaike information criterion (AIC) variable selection method was applied to select appropriate environment variables affecting racing records. The annual improvement of the race time was -0.242 seconds. The model with the lowest AIC value was established when variables were selected in the following order: year, budam classification, jockey ranking, trainer ranking, track condition, weather, age, and gender. The most suitable model was constructed when the jockey ranking and age variables were considered as random effects. Our findings have potential for application as basic data when building models for evaluating genetic abilities of Jeju horses.

The Ecological Values of the Korean Demilitarized Zone(DMZ) and International Natural Protected Areas (비무장지대(DMZ)의 생태적 가치와 국제자연보호지역)

  • Cho, Do-soon
    • Korean Journal of Heritage: History & Science
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    • v.52 no.1
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    • pp.272-287
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    • 2019
  • The Korean Demilitarized Zone (DMZ) was established in 1953 by the Korean War Armistice Agreement. It extends from the estuary of the Imjin River, in the west, to the coast of the East Sea. It is 4 km in width and 148 km in length. However, the ecosystems of the civilian control zone (CCZ) located between the southern border of the DMZ and the civilian control line (CCL) and the CCZ in the estuary of the Han River and the Yellow Sea are similar to those in the DMZ, and, therefore, the ecosystems of the DMZ and the CCZ are collectively known as the "ecosystems of the DMZ and its vicinities." The flora in the DMZ and its vicinities is composed of 1,864 species, which accounts for about 42% of all the vascular plant species on the Korean Peninsula and its affiliated islands. Conducting a detailed survey on the vegetation, flora, and fauna in the DMZ is almost impossible due to the presence of landmines and limitations on the time allowed to be spent in the DMZ. However, to assess the environmental impact of the Munsan-Gaesong railroad reconstruction project, it was possible to undertake a limited vegetation survey within the DMZ in 2001. The vegetation in Jangdan-myeon, in Paju City within the DMZ, was very simple. It was mostly secondary forests dominated by oaks such as Quercus mongolica, Q. acutissima, and Q. variabilis. The other half of the DMZ in Jangdan-myeon was occupied by grassland composed of tall grasses such as Miscanthus sinensis, M. sacchariflorus, and Phragmites japonica. Contrary to the expectation that the DMZ may be covered with pristine mature forests due to more than 60 years of no human interference, the vegetation in the DMZ was composed of simple secondary forests and grasslands formed on former rice paddies and agricultural fields. At present, the only legal protection system planned for the DMZ is the Natural Environment Conservation Act, which ensures that the DMZ would be managed as a nature reserve for only two years following Korean reunification. Therefore, firstly, the DMZ should be designated as a site of domestic legally protected areas such as nature reserve (natural monument), scenic site, national park, etc. In addition, we need to try to designate the DMZ as a UNESCO Biosphere Reserve or as a World Heritage site, or as a Ramsar international wetland for international cooperation. For nomination as a world heritage site, we can emphasize the ecological and landscape value of the wetlands converted from the former rice paddies and the secondary forests maintained by frequent fires initiated by military activities. If the two Koreas unexpectedly reunite without any measures in place for the protection of nature in the DMZ, the conditions prior to the Korean War, such as rice paddies and villages, will return. In order to maintain the current condition of the ecosystems in the DMZ, we have to discuss and prepare for measures including the retention of mines and barbed-wire fences, the construction of roads and railroads in the form of tunnels or bridges, and the maintenance of the current fire regime in the DMZ.