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Development of a Kit for Diagnosing AtCYP78A7 Protein in Abiotic-tolerant Transgenic Rice Overexpressing AtCYP78A7 (AtCYP78A7 과발현 환경스트레스 내성 형질전환 벼의 단백질 진단 키트 개발)

  • Nam, Kyong-Hee;Park, Jung-Ho;Pack, In-Soon;Kim, Ho Bang;Kim, Chang-Gi
    • Journal of Life Science
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    • v.28 no.7
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    • pp.835-840
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    • 2018
  • Quantitative determination of the protein expression levels is one of the most important parts in assessment of the safety of foods derived from genetically modified (GM) crops. Overexpression of AtCYP78A7, a gene encoding cytochrome P450 protein, has been reported to improve tolerance to abiotic stress, such as drought and salt stress, in transgenic rice (Oryza sativa L.). In the present study, an enzyme-linked immunosorbent assay (ELISA) kit for diagnosing AtCYP78A7 protein including AtCYP78A7-specific monoclonal antibody was developed. GST-AtCYP78A7 recombinant protein was induced and purified by affinity column. Four monoclonal antibodies (mAb 6A7, mAb 4C2, mAb 11H6, and mAb 7E8) against recombinant protein were also produced and biotinylated with avidin-HRP. After pairing test using GST-AtCYP78A7 protein and lysate of rice samples, mAb 4C2 and mAb 7E8 were selected as a capture antibody and a detecting antibody, respectively, for ELISA kit. Product test using rice samples indicated that percentages of detected protein in total protein were greater than 0.1% in AtCYP78A7-overexpressing transgenic rice (Line 10B-5 and 18A-4), whereas those in negative control non-transgenic rice (Ilpum and Hwayoung) were less than 0.1%. The ELISA kit developed in this study can be useful for the rapid detection and safety assessment of transgenic rice overexpressing AtCYP78A7.

Detection of Dopamine and Serotonin by Competitive Enzyme-Linked Immunosorbent Assay (경쟁적 ELISA를 이용한 도파민과 세로토닌의 검출)

  • Namkung, Su Min;Choi, Jeong Su;Park, Ji Hyang;Yang, Man Gil;Lee, Min Woo;Kim, Suhng Wook
    • Korean Journal of Clinical Laboratory Science
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    • v.49 no.3
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    • pp.220-226
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    • 2017
  • Dopamine (DA) and serotonin (5-Hydroxytryptamine, 5-HT) are neurotransmitters and hormones that exist in small amounts but have important role in the body. Serum and 24-hour urine are used as specimens, and are usually examined by HPLC-MS. In this study, we tried to detect DA and 5-HT by competitive ELISA using antigen-antibody (Ab) reaction. After immobilizing $5{\mu}g/mL$ BSA conjugate on a 96-well surface, hormone and primary Ab, which are respectively diluted to different concentrations, were treated. Then, HRP-conjugated secondary Ab and TMB were added to measure absorbance. The regression equation and $R^2$ value were calculated based on absorbance, and sensitivity of Ab to hormone as well as the correlation between hormone concentration and absorbance were determined. In DA ELISA, $R^2$, the correlation between the concentration of hormone and absorbance, was the highest by 0.91 when anti-dopamine Ab was diluted 6,000 times and 7,000 times. In 5-HT ELISA, $R^2$ was bigger than 0.90 in every concentration except 3,000 times and 6,000 times. Both DA and 5-HT were not effectively detected at low concentrations (less than $1.0{\times}10^{-7}M$); and because reference value of serum DA is lower than this, HPLC-MS was required to detect serum DA. However, competitive ELISA may be effective in detecting 24-hour urine DA, serum, and 24-hour 5-HT. Further studies are needed to detect hormones more accurately at lower concentrations.

Incidence and Risk Factors for Extended-Spectrum ${\beta}-Lactamase-Producing$ Escherichia coli in Community-acquired Childhood Urinary Tract Infection (지역사회 획득 소아 요로 감염에서 Extended-Spectrum ${\beta}-Lactamase$ 생성)

  • Lee Jung-Won;Shin Jee-Sun;Seo Jeong-Wan;Lee Mi-Ae;Lee Seung-Joo
    • Childhood Kidney Diseases
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    • v.8 no.2
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    • pp.214-222
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    • 2004
  • Purpose: Appropriate antibiotic therapy is important in childhood urinary tract infection and the selection of anibiotics is based on antimicrobial sensitivity of Escherichia coli. Extended-Spectrum ${\beta}-Lactamase(ESBL)$ is an enzyme produced by gram-negative bacilli that has the ability to hydrolyse penicillins, broad-spectrum cephalosporin and monobactam. There have been many reports of outbreaks of hospital infection by ESBL-producing organism. However, community-acquired infection with ESBL-producing organism are rare. This study was performed to retrospectively identify the incidence, characteristics and risk factors of ESBL (+) E. coli in community-acquired childhood UTI. Methods: In 288 children admitted in Ewha Womans University Hospital with E. coli UTI from Mar 2001 to February 2003, ESBL was isolated. ESBL was confirmed by the utilization of an automatized machine(Vitek GNS 433 card) using liquid medium dilution method according to National Committee for Clinical Laboratory Standard. The clinical characteristics, risk factors, antimicrobial resistance and treatment effectiveness were compared with ESBL(-) E. coli UTI. Results: Of 288 E. coli isolates, 31(10.8%) produced ESBL and 93.5%(29/31) occurred in infants younger than 6 month of age(P<0.01). No significant differences were noted in prior antibiotic use, prior admission history and underlying urogenital anomaly. Antimicrobial resistance was significantly higher in ESBL(+) E. coli compared with control patients (P<0.05). Although ceftriaxone showed 100% resistance in ESBL(+) E. coli, bacteriologic sterilization rate after ceftriaxone therapy was higher(96.8%). However, the recurrence rate of febrile UTI within 6 months was higher(25.8%) than control patients(6.6%). Conclusion: Epidemiologic study is required to find out any new risk factors of community-acquired ESBL(+) E. coli UTI and changes in selection of empirical antibiotics should be considered.

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Importance of C-26 Demethylation for Homeostatic Regulation of Brassinosteroids in Seedling Shoots of Zea mays L (옥수수 유식물 신초에서 Brassinosteroids의 항상성 조절을 위반 C-26 탈메틸 반응의 중요성)

  • Park, Hyun-Hee;Kim, Young-Soo;Kim, Seong-Ki
    • Journal of Plant Biotechnology
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    • v.33 no.1
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    • pp.65-73
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    • 2006
  • Regulatory mechanism for endogenous levels of castasterone (CS) and its biosynthetic precursors in shoots of maize was investigated by the use of enzyme solution prepared from the plant tissue. When [$^2H_0$]- and [$^2H_6$]-CS was used as substrates, [$^2H_0$]-26-norCS and [$^2H_3$]-28-norCS were identified as products, indicating that [$^2H_0$]- and [$^2H_6$]-CS are differently metabolized into [$^2H_0$]-26-norCS and [$^2H_3$]-28-norCS by C-26 and C-28 demethylation, respectively. This suggests that both C-26 and C-28 demethylation can be involved in CS catabolism. In fact that C-28 demethylation only occurred when isotope labeled substrate was used, however, C-26 demethylation is thought be a natural reaction occurred in the maize shoots. When 6-deoxoteasterone (6-deoxoTE) was used, 6-deoxo-26-norTE and 3-dehydro-6-deoxo-26-norTE as well as 6-deoxo-3-dehydroTE and 6-deoxotyphasterol (6-deoxoTY) were identified as enzyme products. When 6-deoxoTY was added, 6-deoxo-26-norTY as well as 6-deoxo-3-dehydroTE and 6-deoxoTE was identified as products. These indicate that C-26 demethylation of 6-deoxoTE, 6-deoxo-3-dehydroTE and 6-deoxoTY as well as a reversible C-3 epimerization from 6-deoxoTE to 6-deoxoTY intermediated by 6-deoxo-3-dehydroTE are operative in the maize shoots, demonstrating that endogenous levels of biosynthetic precursors of CS are also controlled by C-26 demethylation. Therefore, it is thought that C-26 demethylation is an important and a common deactivation process which functions to maintain steady state levels of endogenous brassinosteroids in the maize shoots.

Inhibition of Adipocyte Differentiation and Adipogenesis by Aged Black Garlic Extracts in 3T3-L1 Preadipocytes (흑마늘 추출물에 의한 3T3-L1 지방전구세포의 분화 및 adipogenesis 억제에 관한 연구)

  • Park, Jung-Ae;Park, Cheol;Han, Min-Ho;Kim, Byung-Woo;Chung, Yoon-Ho;Choi, Yung-Hyun
    • Journal of Life Science
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    • v.21 no.5
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    • pp.720-728
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    • 2011
  • Garlic (Allium sativum) has been used as a source food as well as a traditional folk medicine ingredient since ancient times. Aged black garlic is a type of fermented garlic and is expected to have stronger anticancer and antioxidant activities than raw garlic. However, the mechanisms of their inhibitory effects on adipocyte differentiation and adipogenesis are poorly understood. In the present study, the effects and mechanisms of water extracts of raw garlic (WERG) and aged black garlic (WEABG) on adipocyte differentiation and adipogenesis in 3T3-L1 preadipocytes were investigated. Treatment with WEABG significantly suppressed terminal differentiation of 3T3-L1 preadipocytes in a dose-dependent manner as confirmed by a decrease in lipid droplet number and lipid content through Oil Red O staining, however WERG had no such effect. In addition, WEABG reduced accumulation of cellular triglyceride, which is associated with a significant inhibition of key pro-adipogenic transcription factors including peroxisome proliferator-activated receptor ${\gamma}$ (PPAR${\gamma}$), cytidine-cytidine-adenosine-adenosine-thymidine (CCAAT)/enhancer binding proteins ${\alpha}$ (C/EBP${\alpha}$) and C/EBP${\beta}$. Taken together, these results provide important new insight that aged black garlic might inhibit adipogenesis by suppressing the pro-adipogenic transcription factors in 3T3-L1 preadipocytes, and further studies will be needed to identify the active compounds that confer the anti-obesity activity of aged black garlic.

Galactosemia Detected by Neonatal Screening Test (신생아 선별검사에 의해 발견된 갈락토스혈증에 대한 고찰)

  • Park, Il Sung;Cho, Hye Jung;Lee, Dong Hwan;Song, Jung Hwan
    • Clinical and Experimental Pediatrics
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    • v.46 no.5
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    • pp.440-446
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    • 2003
  • Purpose : The genetic disturbance of galactosemia is expressed as a cellular deficiency of either galactose-1-phosphate uridyltransferase(GALT) or galactokinase(GALK) or UDP galactose 4-epimerase(GALE). To find-out the pattern of galactosemia in Korea, we retrospectively analyzed cases of galactosemia detected by neonatal screening program. Methods : We analyzed medical records of patients who visited Soonchunhyang University Hospital at age of 1 month after showing abnormalities in neonatal screening of galactosemia. For accurate diagnosis, galactose was measured by enzyme immunoassay(EIA) and fluorophotometer, also galactose-1-phosphate by fluorophotometer. Enzyme activities of GALK, GALT and GALE in RBC and galactose-1-phosphate were measured by radioisotope assay(RIA). Beutler test were done. Patients went on a lactose-free diet and follow-up tests for galactose, galactose-1-phosphate level and enzyme activity were performed. Results : 10 patients(male : 6, female : 4) were diagnosed as galactosemia. Two patients had GALK deficiency and two had GALT deficiency. Six were GALE deficient showing the largest number. In two patients with GALK deficiency, GALT and GALE activities were normal but GALK activities showed respectively reduced activity. For GALT deficiency, two patients had low GALT activity in RBC and showed genotype of Duarte 2/G(galactosemia) in DNA analysis. In one patient, GALT activity was normal. Three patients seemed to be heterozygote state of GALE deficiency according to GALE activity levels. Four patients showed GALK hyperactivity. Conclusion : GALE deficiency provided the highest number. After lactose-free diet, galactose and galactose-1-phosphate were normaly maintained. Neonatal screening on galactosemia is essential for preventing life-threatening symptoms and an accurate diagnosis is needed for finding out the type of galactosemia which is important for prognosis.

Research of z-axis geometric dose efficiency in multi-detector computed tomography (MDCT 장치의 z-축 기하학적 선량효율에 관한 연구)

  • Kim, You-Hyun;Kim, Moon-Chan
    • Journal of radiological science and technology
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    • v.29 no.3
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    • pp.167-175
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    • 2006
  • With the recent prevalence of helical CT and multi-slice CT, which deliver higher radiation dose than conventional CT due to overbeaming effect in X-ray exposure and interpolation technique in image reconstruction. Although multi-detector and helical CT scanner provide a variety of opportunities for patient dose reduction, the potential risk for high radiation levels in CT examination can't be overemphasized in spite of acquiring more diagnostic information. So much more concerns is necessary about dose characteristics of CT scanner, especially dose efficient design as well as dose modulation software, because dose efficiency built into the scanner's design is probably the most important aspect of successful low dose clinical performance. This study was conducted to evaluate z-axis geometric dose efficiency in single detector CT and each level multi-detector CT, as well as to compare z-axis dose efficiency with change of technical scan parameters such as focal spot size of tube, beam collimation, detector combination, scan mode, pitch size, slice width and interval. The results obtained were as follows ; 1. SDCT was most highest and 4 MDCT was most lowest in z-axis geometric dose efficiency among SDCT, 4, 8, 16, 64 slice MDCT made by GE manufacture. 2. Small focal spot was 0.67-13.62% higher than large focal spot in z-axis geometric dose efficiency at MDCT. 3. Large beam collimation was 3.13-51.52% higher than small beam collimation in z-axis geometric dose efficiency at MDCT. 4. Z-axis geometric dose efficiency was same at 4 slice MDCT in all condition and 8 slice MDCT of large beam collimation with change of detector combination, but was changed irregularly at 8 slice MDCT of small beam collimation and 16 slice MDCT in all condition with change of detector combination. 5. There was no significant difference for z-axis geometric dose efficiency between conventional scan and helical scan, and with change of pitch factor, as well as change of slice width or interval for image reconstruction. As a conclusion, for reduction of patient radiation dose delivered from CT examination we are particularly concerned with dose efficiency of equipment and have to select proper scanning parameters which increase z-axis geometric dose efficiency within the range of preserving optimum clinical information in MDCT examination.

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Monitoring of Chemical Properties from Paddy Soil in Gyeongnam Province (경남지역 논 토양 화학성분 변동조사)

  • Lee, Young-Han;Lee, Seong-Tae;Heo, Jae-Young;Kim, Min-Geun;Hong, Kang-Pyo;Song, Won-Doo;Rho, Chi-Woong;Lee, Jin-Ho;Jeon, Weon-Tai;Ko, Byong-Gu;Roh, Kee-An;Ha, Sang-Keun
    • Korean Journal of Soil Science and Fertilizer
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    • v.43 no.2
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    • pp.140-146
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    • 2010
  • Monitoring of the dynamic changes at paddy rice agriculture is very important for agricultural sustainability. Field monitoring was performed to evaluate the soil chemical properties of 260 paddy soil samples every four years from 1999 to 2007 in Gyeongnam Province. Soil chemical properties such as pH, organic matter, available phosphate, silicate, exchangeable potassium, calcium, and magnesium contents were analyzed. The contents of exchangeable cations, and available silicate were significantly increased in 2007 compared to 1999. The chemical contents of organic matter, exchangeable potassium, and magnesium were significantly increased in acid sulfate soil, and silty clay loam compared to those of other soil types, and textures. Especially, content of organic matter was significantly increased in hill area compared to other soil topographies, while exchangeable potassium was significantly decreased. Principle component analysis (PCA) of chemical properties in paddy soils was obtained with eigenvalues > 1 summing 39.1%of variance for PC1, 20.4%of variance for PC2, and 59.5%of the total variance in the all of soil chemical properties. Therefore, principal component analysis is more effective for monitoring from chemical properties of paddy soil.

EST Profiling for Seed-hair Characteristic and Development of EST-SSR and SNP Markers in Carrot (당근 종모 형질 관련 EST profiling과 이를 이용한 EST-SSR 및 SNP 마커 개발)

  • Oh, Gyu-Dong;Hwang, Eun-Mi;Shim, Eun-Jo;Jeon, Sang-Jin;Park, Young-Doo
    • Horticultural Science & Technology
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    • v.28 no.6
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    • pp.1025-1038
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    • 2010
  • Carrot ($Daucus$ $carota$ L. var. $sativa$) is one of the most widely used crops in the world. Moreover it is an important crop because of its high content of ${\beta}$-carotene, well-known as the precursor of vitamin A carotenoid. However, seed-hair which is generated in epidermal cell of seeds inhibits absorption and germination. For that reason, carrot seeds are commercialized after mechanical hair removal process. To overcome such cumbersome weaknesses, new breeding program for developing hairless-seed carrot cultivar has been needed. Therefore, in this study, cDNA libraries from seeds of short-hair seed phenotype CT-ATR615 OP 666-13line and hairy seed CT-ATR615 OP-CK1-9 line were constructed and expression patterns related to generation of seed-hair were analyzed by comparison of EST sequences. Differential EST sequence results between two lines were classified into FunCat functional categories based on the results of BlastX search. Higher expression quantities belonging to metabolic category were shown on short-hair seed line than hairy-seed one. Differential expression quantities between those two lines in the protein folding and stabilization, subcellular localization categories were supposed to contribute variously on the generation of seed-hair. We confirmed 50 and 59 SSR sites, and 2 SNP sites by analyzing EST sequences in two lines; thereafter, we designed SNP and SSR primer sets from these EST sequence information as a molecular marker. These markers are thought to be used in research of molecular markers for classification of carrot family and related to various traits, as well as seed-hair characteristic.

Clinical Characteristics of Patients with Traumatic Diaphragm Injury and Comparison of Scoring Systems as Predictors (외상성 횡격막 손상환자의 임상적 고찰과 예후 예측 인자의 유용성 비교)

  • Lee Sang Jin;Jung Jin Hee;Sohn Dong Suep;Cho Dai Yun
    • Journal of Chest Surgery
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    • v.38 no.1 s.246
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    • pp.56-62
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    • 2005
  • Because of high morbidity and mortality, traumatic diaphragm injury remains a diagnostic challenge. In this study, we evaluate that some factors and scores can be used as predictors. Material and Method: From May 1995 to June 2003, 23 patients with traumatic diaphragm injury were enrolled. We examined the clinical features of patients. RTS, TRISS, ISS and APACHE II scores for each patient are calculated for analyzing the relationship of mortality and ICU duration. Result: The study identified 15 men $(65.2{\%})$ and 8women $(34.8{\%})$. There are right sided diaphragmatic injury in 11 patients $(47.8{\%})$, left sided in 11 $(47.8{\%})$, and both sided in 1 ($0.4{\%}$). Plain X-ray, CT, upper GI contrast study and esophagogastroscopy were used as diagnostic tools. Age, hemodynamic status, early diagnosis are not associated with outcome. As prognostic factor, RTS and ISS are associated with mortality and there was negative relationship between RTS and ICU duration (r=0.737, p=0.026). Conclusion: An early diagnosis of traumatic diaphragm injury can frequently be missed in the acute trauma setting. So high index of suspicion and a careful examination are important in multiple trauma patients. An RTS can probably be used effectively as a predictor for the severity and prognosis in patients with traumatic diaphragm injury.