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식품과 알레르기: 유전자 재조합 식품의 알레르기 위험성

  • 손대열
    • Proceedings of the Korean Journal of Food and Nutrition Conference
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    • 2000.12a
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    • pp.29-34
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    • 2000
  • 산업 발달에 따라 날로 많은 식품들이 새롭게 개발되어지고 있다. 또한 이와 병행해서 식품으로 인한 알레르기 발생 빈도도 날로 증가하고 있으며 그 증상 또한 점차 심화되고 있는 것이 세계적인 추세이다. 우리나라도 예외는 아니어서 일반 알레르기 환자뿐 아니라 식품으로 인한 알레르기 환자들이 점차 증가됨이 보고되어지고 있다. 농산물 시장의 수입개방이후 우리나라에는 많은 해외 농산물이 수입되어지고 있으며 그 중 작년 한해의 경우 총 수입 농산물의 10%를 넘는 유전자 재조합 농산물이 우리나라에 수입되어진 것으로 통계 보고되어졌다. 이러한 관점에서 알레르기 환자의 증가와 새로운 식품 (특히 유전자 재조합 식품)의 증가에는 서로 관련성이 있을 것으로 추측되어지고 있어 (새로운) 식품에 대한 알레르기성의 예측과 관리가 필요한 실정이다. 이에 몇몇 발표된 유전자 재조합 식품에 관련된 알레르기성 검사 논문들과 실험실에서 이루어진 연구 결과들을 중심으로 유전자 재조합 식품의 알레르기 위험성에 대해 알아보고자 한다. 일반적으로 식품의 단백질이 알레르겐(allergen)으로 작용하기 위해서는 먼저 소화효소에 의해 분해되어지고 장에서 흡수되어져서 immunopotent cell에 의해 process 되어 immune system에 present 되어져야 한다. 따라서 단백질로 인한 알레르기 반응은 그 단백질의 자연적 형태 뿐만이 아니라 소화 효소에 분해된 단편들의 구조 또는 다른 알레르겐 단백질과의 유사 구조로 인한 교차 반응에 의해 발생함을 기억해야 한다. 식품 단백질 중 어떤 단백질이 알레르겐으로 작용하는가에 대한 특이성 조사에 많은 관심이 집중되어지고 있지만 아직까지는 대략 다섯 개 정도의 일반적인 특성으로서 요약되어질 수 있다. 그러나 이러한 대략의 특성에 적용되지 않는 식품 알레르겐도 많음을 잊어서는 안 될 것이다. 알레르겐으로 작용하는 식품 단백질의 일반적 특성 1. 좋은 수용성 2. 식품내에 많은 부분을 차지하는 주 단백질이 주 알레르겐으로 작용 3. 단백질내에 하나 이상의 IgE-binding site 존재 4. 위장액에 대한 저항성 5. 10~70 kDa 크기 유전자 재조합 기술이란 말 그대로 유전자를 인위적으로 새롭게 조합하는 기술로 이전의 기술로는 불가능했던 유전적 변형을 농작물과 동물에 가능하게 했으며 이로 인해 유전적으로 변형된 식용 동식물의 개발이 가능하게 되었다. 새로운 유전인자를 개체에 삽입함으로 새로운 단백질이 발현 될수 있고 그로 인해 1) 해충과 질병에 대한 저항성 증가, 2) 화학 제초제에 대한 새로운 저항성 부여, 3) 식품의 저장성 향상, 4) 식품의 영향적 보충/향상 등의 이점을 얻을 수 있다 (표 1). 세계적으로 유전자 재조합 된 새로운 농산물의 재배는 날로 증가추세에 있으며 그 중에서 가장 많은 부분을 차지하는 농산물로 soybean을 들 수 있으며 (표 2) soybean을 중심으로 그 알레르기성의 변화가 연구 조사된 몇 가지 예를 살펴보고자 한다. (표 3)에 요약된 soybean중 첫 번째 경우는 재초제에 대한 저항성을 높여주기 위해 Agrobacterium에 존재하는 EPSPS라는 단백질을 콩에서 발현하도록 찬 유전자 재조합 된 콩의 경우이다. 이 콩의 경우에는 첫째. 이전된 새로운 단백질 EPSPS가 다른 여러 식물에 이미 존재하고 있는 단백질로서 우리가 이미 이러한 식품을 섭취할 때 이 단백질도 같이 섭취해오고 있었다는 점, 둘째. 이 단백질이 소화액 분해 실험에서 짧은 시간내에 분해가 되었다는 점, 셋째. 재조합 된 콩과 자연 콩이 성분 분석에서 차이를 나타내지 않았다는 점, 네 번째. 쥐를 통한 다양섭취 실험에서 아무런 이상 반응이 없었다는 점등의 결과를 기준으로 알레르기에 대한 개별 검사 없이 안전한 콩으로 결론짓고 있다. 영양성을 높이기 위해 Brazil nut에서 methionine 함량이 풍부한 2s albumine을 콩에서 발현하도록 한 두 번째 유전자 재조합 콩의 경우 이전된 단백질 때문에 Brazil nut에 알레르기 반응을 일으키는 알레르기 환자들을 조사한 결과 역시 재조합 된 콩에도 알레르기 반응을 일으켰다는 보고이다. Brazil nut에서 콩으로 이전된 단백질이 Brazil nut에서의 알레르기성을 그대로 유지한 점을 볼 때 새로운 단백질이 어디에서 유래하는가가 중요함을 잘 보여준 연구이다 세 번째 콩의 경우 역시 영양성을 높여주기 위해 corn에서 10 kDa과 HSZ 단백질을 콩에서 발현하도록 유전자 재조합했는데 이 콩의 경우는 알레르기 환자들이 유전자 재조합 된 콩과 자연 콩에 반응의 차이를 나타내지 않았다는 결과 보고이다. 위의 세 실험 결과들을 종합해 볼 때 무엇보다도 새롭게 발현된 단백질이 원래 어떤 성질을 갖고 있으며 어디에서 유래했는지가 알레르기성 조사에 중요한 역할을 한다 할 수 있겠다. 또한 유전자 재조합된 식품들은 알레르기 환자들을 위해 표기되어져야 할 것인데 이를 위한 알레르기성 검사 실험은 공공단체를 통해 이루어져야 할 것이며 환자들마다 알레르겐으로 작용하는 단백질의 인식부위(epitope)가 다를 수 있기 때문에 적어도 10명 이상의 알레르기 환자들이 조사되어져서 검사가 이루어져야 할 것이다. 환자들의 혈청을 통한 in vitro 실험에서는 ELISA, RAST, immunoblotting과 같은 검사 방법들이 적용될 수 있고, 그 결과가 음성인 경우에 그 다음 단계로 in vivo 실험에서는 직접 환자의 피부반응검사 (skin prick test)나 DBPCFC (double-blind placebo-controlled food challenge) 검사 방법을 통해 확인되어져서 이 모든 경우가 음성인 경우와 하나라도 양성인 경우를 구별하여 식품에 표기함으로 알레르기 환자들의 유전자 재조합 식품에 대한 안전성이 보장되어져야 할 것이다.

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Influence of Water Foxtail on Growth of Rice and Weed in No-Tillage Transplanted Rice (벼 무경운재배시 둑새풀 우점 정도가 벼 생육 및 잡초 발생에 미치는 영향)

  • 강양순;곽태순;송문태
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.41 no.5
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    • pp.619-624
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    • 1996
  • An experiment was carried out to find out the effects of water foxtail on weed control and rice yield as well as on the reduction of nitrogen application in no tillage transplanted paddy field. Paddy field was dominated by water foxtail whose soil covering degree was adjusted from 8 to 6 by treatment of paraquat (70 l /10a of solution diluted by 1, 000 times). Thirty five-day old seedling was mechanically transplanted and reduced nitrogen, 80% of conventional application was applied at the paddy field. The higher failure in seedling stand was observed at higher degree of soil covering by water foxtail. The failure of seedling stand with covering degree of 8(no paraquat) was 37.4%, while that of covering degree of 6(paraquat-treated) was 12.3%. However, the seedling stand failure was completely recovered at covering degree of six at two weeks after transplanting. The mechanical transplanting made water foxtail in the paddy field fall on the ground whose panicle part was recovered from falling sometimes after transplanting and whole plant died with slow scenesence untill the late of June. But the water foxtail affected by paraquat produced the new panicle from uppernode of stem with dead leaves at early of June and it become die slowly untill the early of July. Though several rice field weeds were observed in the order of Echinochloa crusgallis, Cyperus serotinus, Lemma paucicostata during the active tillering stage of rice, the value of weed control due to the dominance of water foxtail was 77~78%. The death and subsequent decay of water foxtail during the active tillering stage of rice induced the soil reduction which again defect the growth of rice root. The more vigerous rice growth was observed in the plot dominated with water foxtails, than control plot. The yields of rice in the water foxtail (degree 6) plot was 629kg per l0a as brown rice, the same as that of control plot. In conclusion, the no-tillage transplanting with control of covering degree of water foxtail (to degree 6) was effective in reduction of input cost such as herbicide and nitrogen fertilizer, as well as weed control without reduction of rice yield.

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Effects of Corticosteroid on the Paraquat Induced Lung Injury (Paraquat에 의한 급성 폐손상에 대한 Corticosteroid의 영향)

  • Chang, Keun;Kim, An-Myung;Kang, Jeong-Seong;Jung, Byung-Hak;Jeong, Eun-Taik;Moon, Hyung-Bae
    • Tuberculosis and Respiratory Diseases
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    • v.39 no.4
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    • pp.325-333
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    • 1992
  • Background and Methods: To study the effects of corticosteroid (CS) on the parquat (PQ) induced lung injury, serial cellular analyses of bronchoalveolar lavage (BAL) fluid were done with simultaneous histopathologic examination after intraperitoneal injection of PQ on the rats. The sacrificed animals were divided into three groups; control group, PQ group received intraperitoneal injection of 20 mg/kg of PQ, and CS group received daily injection of Methylprednisolone sodium succinate (20 mg/kg) in addition to PQ. Results: 1) Cellular analyses of BAL fluid: The total cell count in the BAL fluid were increased gradually from 6 hours after PQ administration (p<0.05), and was decreased at 3 days after (p<0.05). These changes were mainly due to the effects of PQ on the neutrophil influx (p<0.05). But, the number of macrophage and the percentage of lymphocyte in total cells showed little changes. The CS administration showed the suppression of neutrophil influx in the BAL fluid (p<0.05), but could not show any significant effect on the number of macrophage and lymphocyte. 2) Histopathologic examination: In the PQ group, inflammatory changes especially with prominant neutrophil infiltration were gradually progressed over time. Those changes were found in both alveolar space and interstitium with resultant alveolar structural changes, but subsided from 3 days after. CS suppressed inflammatory changes in the alvolar space and interstitium, especially with decreased infiltration of neutrophil. Conclusion: CS suppressed neutrophil infiltration in the acute lung injury induced by PQ, those findings were ascertained by serial cellular analyses of BAL fluid and histopathologic examination.

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Analysis of Methoxyfenozide and Bentazone in Plant with HPLC by pH-Adjusted Liquid-liquid Partition Cleanup (pH조절 분배추출에 의한 식물체 중 Methoxyfenozide와 Bentazone의 HPLC 분석)

  • Lo, Seog-Cho;Hwang, Cheol-Hwan;Yoo, Ki-Yong;Han, Seong-Soo
    • Korean Journal of Environmental Agriculture
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    • v.26 no.3
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    • pp.246-253
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    • 2007
  • This study was performed to enhance the cleanup efficiency of methoxyfenozide and bentazone by pH adjustment in the course of liquid-liquid partition and to develop an optimum analytical conditions using HPLC coupled with DAD for two matrices, brown rice and rice straw. Preparation procedure of brown rice sample was "extraction${\rightarrow}$coagulation${\rightarrow}$liquid-liquid partition$\rightarrow$-florisil C.C", and this procedure was samely applied to two compounds. In rice straw, preparation procedure of methoxyfenozide sample was "extraction$\rightarrow$-alkalization$\rightarrow$liquid-liquid extraction$\rightarrow$coagulation$\rightarrow$florisil C.C", and in the case of bentazone, "extraction$\rightarrow$alkalization$\rightarrow$liquid-liquid partition$\rightarrow$acidification$\rightarrow$liquid-liquid extraction$\rightarrow$florisil C.C". All these purified samples were redissolved in the mobile phases, acetonitile : 20 mM sodium acetate (75:25, v/v) for methoxyfenozide and acetonitrile : 75 mM sodium acetate, pH 6.0 (40:60, v/v) for bentazone. Recoveries of methoxyfenozide analysis in brown rice and rice straw were 83.5-97.4 and 86.4-97.3%, and detection limits were 0.02 and 0.04 mg/kg, respectively. Recoveries of bentazone in brown rice and rice straw were 86.8-101.9 and 88.3-94.5% and detection limits were 0.005 and 0.01 mg/kg, respectively. This methods seem to be usefully applied to the residue analysis of two compounds in the view of producing stable analytical condition and fair reproducibility.

The Construction and Management of Artificial Wetland Using Emergent Macrophytes for High Biomass Production (대형정수식물을 활용한 높은 생산성의 인공습지 조성 및 관리)

  • Hong, Mun Gi;Heo, Young Jin;Kim, Jae Geun
    • Journal of Wetlands Research
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    • v.16 no.1
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    • pp.61-72
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    • 2014
  • To present a guideline on the construction and management of artificial wetlands for high biomass production, three emergent macrophytes (Phragmites australis, PA; Typha angustifolia, TA; and Zizania latifolia, ZL) were planted under two substrates conditions (general soil with and without moss peat) and two water levels (5 cm and 20 cm) and monitored for three years. ZL showed greater growth performance rather than the others not only at early growth phase in the first year [shoot height, 200 cm; above-ground dry weight (AGDW), 500 $g/m^2$] but also in the last year (ZL, 1,100 $g/m^2$; TA, 770 $g/m^2$; and PA, 450 $g/m^2$ of AGDW). ZL with rapid growth at the early growth phase was not affected by naturally introduced weeds, whereas slower and poorer growth of PA and TA at the early growth phase resulted in relatively higher introduction and establishment of natural weeds. In turn, such introduced weeds negatively contributed to the growth of PA and TA particularly under shallow water (5 cm) with the substrate condition including moss peat. We suggest a plant material with rapid and great growth at the early phase such as ZL for reducing possible negative influences by the natural weeds and wild animals for high biomass production in constructed wetlands. A pre-growing process in greenhouse prior to planting might be an useful option to raise the competitiveness of those species when planting PA and/or TA. In addition, we recommend that integrated weed management system with utilizing various options at the most appropriate timing must be applied for maintaining sustainable high biomass production at the artificial wetlands.

Volatilization of molinate in paddy rice ecosystem and its concentration in air causing phytotoxicity to chili pepper (벼 재배 환경 중 molinate의 휘산과 공기 중 고추약해 발현농도)

  • Park, Byung-Jun;Choi, Ju-Hyeon;Kim, Chan-Sub;Im, Geon-Jae;Oh, Byung-Youl;Shim, Jae-Han
    • The Korean Journal of Pesticide Science
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    • v.9 no.1
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    • pp.70-80
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    • 2005
  • To evaluate the exposure of molinate in agricultural environment and its effect against the non-target crop in air, this experiment was conducted to elucidate volatilization characteristics of molinate in aquatic condition and to determine critical concentration of molinate in the air causing phytotoxicity to Chili pepper. Cumulative volatilized rate of molinate from water was 22.7% at $35^{\circ}C$ for water temperature and 20 L/min for air velocity while 3.2% at $25^{\circ}C$ and 10 L/min within 47 hour after applied under closed system, respectively. The molinate concentrations in air above 60 cm height from soil surface of valley and open paddy rice field were reached the highest value of 18.17 and $11.59{\mu}g/m^3$, respectively within 24 hours after applying granular formulation at dose rate of molinate 150 g/1,000 $m^2$. However, their concentrations were drastically diminished to around 0.18 and $0.51{\mu}g/m^3$ level in 20 days after application, which volatilization pattern were similar to both regions. Also, the concentration of molinate in air above 60 cm height from soil surface was distributed higher 2 times than that above 180 cm height. Meanwhile, a phytotoxic symptom against the nearby chili pepper was revealed within three days after applied and molinate was detected $0.004{\sim}0.006$ mg/kg level from severe damaged leaves. The dose and exposure relations of molinate in the air against the non-target crop was also investigated in lab trial. The phytotoxic symptom, shriveled leaves, of the chili pepper was encountered by exposing two days with concentration of $13.6{\mu}g/m^3$, three days with $6.8{\mu}g/m^3$ or four days with $3.4{\mu}g/m^3$. The symptom was still recovered within four weeks after the plants had received fresh air. On the other hand, the phytotoxic response through root uptake of the herbicide in water culture was relatively insensitive, in which the symptom is observed ten days with the concentration of 300 ${\mu}g/L$.

Effects of Aminotriazole on Lung Toxicity of Paraquat Intoxicated Mice (Paraquat중독에 의한 폐독성에 미치는 Aminotriazole의 영향)

  • Lee, Seung-Il;An, Gi-Wan;Chung, Choon-Hae
    • Tuberculosis and Respiratory Diseases
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    • v.41 no.3
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    • pp.222-230
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    • 1994
  • Background: Paraquat, a widely used herbicide, is extremely toxic, causing multiple organ failure in humans. Paraquat especially leads to irreversible progressive pulmonary fibrosis, which is related to oxygen free radicals. However, its biochemical mechanism is not clear. Natural mechanisms that prevent damage from oxygen free radicals include changes in glutathione level, G6PDH, superoxide dismutase(SOD), catalase, and glutathione peroxidase. The authors think catalase is closely related to paraquat toxicity in the lungs Method: The effects of 3-amino-1,2,4-triazole(aminotriazole), a catalase inhibitor, on mice administered with paraquat were investigated. We studied the effects of aminotriazole on the survival of mice administered with paraquat, by comparing life spans between the group to which paraquat had been administered and the group to which a combination of paraquat and aminotriazole had been administered. We measured glutathion level, glucose 6-phosphate dehydrogenase(G6PDH), superoxide dismutase(SOD), catalase, and glutathione peroxidase(GPx) in the lung tissue of 4 groups of mice: the control group, group A(aminotriazole injected), group B(paraquat administered), group C(paraquat and aminotriazole administered). Results: The mortality of mice administered with paraquat which were treated with aminotriazole was significantly increased compared with those of mice not treated with aminotriazole. Glutathione level in group B was decreased by 20%, a significant decrease compared with the control group. However, this level was not changed by the administration of aminotriazole(group C). The activity of G6PDH in all groups was not significantly changed compared with the control group. The activities of SOD, catalase, and glutathione peroxidase(GPx) in the lung tissue were significantly decreased by paraquat administration(group B); catalase showed the largest decrease. Catalase and GPX were significantly decreased by aminotriazole treatment in mice administered with paraquat but change in SOD activity was not significant(group C). Conclusion: Decrease in catalase activity by paraquat suggests that paraquat toxicity in the lungs is closely related to catalase activity. Paraquat toxicity in mice is enhanced by aminotriazole administration, and its result is related to the decrease of catalase activity rather than glutathione level in the lungs. Production of hydroxyl radicals, the most reactive oxygen metabolite, is accelerated due to increased hydrogen peroxide by catalase inhibition and the lung damage probably results from nonspecific tissue injury of hydroxyl radicals.

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Historical Change of Population Abundances of Panonychus ulmi and Tetranychus urticae (Acari: Tetranychidae) in Selected Apple Orchards in Suwon and Its Hypothetical Explanation (수원 지역 사과원에서 사과응애와 점박이응애 개체군의 역사적 변천과정 및 해석)

  • Kim Dong Soon;Lee Joon Ho
    • Korean journal of applied entomology
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    • v.44 no.2
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    • pp.115-123
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    • 2005
  • Historical changes of population abundances of European red mite (ERM), Panonychus ulmi (Koch), and two-spotted spider mite (TSSM), Tetranychus urticae (Koch) (Acari: Tetranychidae), were described in selected apple orchards in the National Horticultural Research Institute (NHRI, Suwon, Korea), based on research reports of the NHRI from 1958 to 1998. ERM was an abundant species up to 1970, and TSSM became a dominant species after 1980. The change occurred around mid 1970. Three hypotheses were made to explain the change: TSSM competitively replaces ERM, ground cover weeds are a major influencing factor on movement of TSSM (TSSM movement into trees is accelerated by destroying weeds), and ERM and TSSM populations are regulated by natural enemy complexes when the orchard system is not disrupted. And long-term results of the interaction between two species were projected according to the combination of different orchard management strategies: pesticide sprays (non-selective toxic pesticide spray : heavy pesticide pressure (HPP), and selective soft pesticide spray = low pesticide pressure (LPP)) and weed control methods (grass planting, and clean culture system with herbicides). In the HPP and grass planting system, ERMs are abundant because ERM can avoid competition with TSSM as movement of TSSM to trees are restricted, and natural enemy complexes are destroyed by toxic pesticides. In the HPP and clean culture system, TSSMs are abundant because TSSM moves to trees from early season and competitively replaces ERM. In the LPP and grass planting system, ERMs are abundant because movement of TSSM to trees is reduced, but they do not build up a high population density since their densities are regulated by natural enemy complexes. In the LPP and clean culture system, TSSM moves to trees and competes with ERM, but the competition pressure is reduced because population densities of mites are regulated in a lower level by natural enemy complexes. So, ERM can occurs in late season. Thus, two species can coexist temporarily with more ERM in early season and more TSSM in late season. TSSM abundant phenomenon presented in this study can be partially explained as a result of long-term interaction between ERM and TSSM under the HPP and clean culture system.

Comparison of Acute Symptoms Between a Alpine Agricultural Workers and General Workers in Gangwon-do (강원도 일부지역 고랭지 농업 농민과 일반 농업 농민의 농약중독 증상 비교)

  • Song, Jae-Seok;Park, Woong-Sub;Kwak, Yeon-Hee;Seo, Jong-Chul;Choi, Hong-Soon
    • Journal of agricultural medicine and community health
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    • v.28 no.1
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    • pp.29-37
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    • 2003
  • Objectives: This study was performed to evaluate the pesticide exposure status and acute pesticide poisoning symtoms among agricultural workers at Gangwon-do province alpine area. Alpine area was defined as the area higher than 400m. Methods: We analyzed 257 interviwed questionnaire about pesticide exposure, acute pesticide poisoning symptoms and other variables. Results: The result was shown that agricultural workers at alpine area used more pesticide than general agricultural workers for annual usage days(35.9 days vs 14.4 days), daily usage hours(6.7 hrs vs 2.8hrs, p<0.05). But there was no difference between general and alpine agricultural worker's herbicide exposure. Moreover, the alpine agricultural worker's acute pesticide poisoning symtom score was higher than general agricultural worker's. These difference was also found at result of regression analysis, under control the age, sex, monthly income. Most frequently suffered symptom was headache and dermatological problems. The symptom prevalence of dermatological problem, headache, general weakness, eye irritation, nausea were higher among alpine agricultural workers than general agricultural workers. Conclusions: As a result, agricultural workers at alpine area were more exposed to pesticide and suffered from pesticide poisoning symptoms. To prevent the symptoms and disease from pesticide exposure among agricultural workers at alpine area, more research and political effort will be needed.

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Generation of Transgenic Rice without Antibiotic Selection Marker through Agrobacterium-mediated Co-transformation System (아그로박테리움 동시 형질전환 시스템을 통한 항생제 선발 마커가 없는 형질전환벼의 생산)

  • Park, Soo-Kwon;Kwon, Tack-Min;Lee, Jong-Hee;Shin, Dong-Jin;Hwang, Woon-Ha;Song, You-Chun;Cho, Jun-Hyun;Nam, Min-Hee;Jeon, Seung-Ho;Lee, Sang-Yeol;Park, Dong-Soo
    • Journal of Life Science
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    • v.22 no.9
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    • pp.1152-1158
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    • 2012
  • Development of transgenic plant increasing crop yield or disease resistance is good way to solve the world food shortage. However, the persistence of marker genes in crops leads to serious public concerns about the safety of transgenic crops. In the present paper, we developed marker-free transgenic rice inserted high molecular-weight glutenin subunit (HMW-GS) gene ($D{\times}5$) from the Korean wheat cultivar 'Jokyeong' using Agrobacterium-mediated co-transformation method. Two expression cassettes comprised of separate DNA fragments containing only the $D{\times}5$ and hygromycin resistance (HPTII) genes were introduced separately into Agrobacterium tumefaciens EHA105 strain for co-infection. Each EHA105 strain harboring $D{\times}5$ or HPTII was infected into rice calli at a 3: 1 ratio of EHA105 with $D{\times}5$ gene and EHA105 with HPTII gene expressing cassette. Then, among 66 hygromycin-resistant transformants, we obtained two transgenic lines inserted with both the $D{\times}5$ and HPTII genes into the rice genome. We reconfirmed integration of the $D{\times}5$ and HPTII genes into the rice genome by Southern blot analysis. Wheat $D{\times}5$ transcripts in $T_1$ rice seeds were examined with semi-quantitative RT-PCR. Finally, the marker-free plants containing only the $D{\times}5$ gene were successfully screened at the $T_1$ generation. These results show that a co-infection system with two expression cassettes could be an efficient strategy to generate marker-free transgenic rice plants.