• Title/Summary/Keyword: Screening system

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Identification of HUGT1 as a Potential BiP Activator and a Cellular Target for Improvement of Recombinant Protein Production Using a cDNA Screening System

  • Ku, Sebastian Chih Yuan;Lwa, Teng Rhui;Giam, Maybelline;Yap, Miranda Gek Sim;Chao, Sheng-Hao
    • Molecules and Cells
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    • v.27 no.5
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    • pp.577-582
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    • 2009
  • The development of a high-throughput functional genomic screening provides a novel and expeditious approach in identifying critical genes involved in specific biological processes. Here we describe a cell-based cDNA screening system to identify the transcription activators of BiP, an endoplasmic reticulum (ER) chaperone protein. BiP promoter contains the ER stress element which is commonly present in the genes involved in unfolded protein response (UPR) that regulates protein secretion in cells. Therefore, the positive regulators of BiP may also be utilized to improve the recombinant protein production through modulation of UPR. Four BiP activators, including human UDP-glucose:glycoprotein glucosyltransferase 1 (HUGT1), are identified by the cDNA screening. Overexpression of HUGT1 leads to a significant increase in the production of recombinant erythropoietin, interferon ${\gamma}$, and monoclonal antibody in HEK293 cells. Our results demonstrate that the cDNA screening for BiP activators may be effective to identify the novel BiP regulators and HUGT1 may serve as an ideal target gene for improving the recombinant protein production in mammalian cells.

High Throughput Screening for Searching a New Inhibitors of Acetolactate Synthase (Acetolactate synthase에 대한 고효율 활성 측정방법 및 신규 저해제 탐색)

  • Park, S.H.;Lee, K.H.;Choi, J.S.;Pyon, J.Y.;Cho, K.Y.;Hwang, I.T.
    • The Korean Journal of Pesticide Science
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    • v.5 no.3
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    • pp.41-46
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    • 2001
  • This study was conducted to develop a high throughput system for screening acetolactate synthase(ALS) inhibitors, and to detect basic mother molecules for developing new novel herbicide candidates. The high throughput screening (HTS) method using 96-well plate and microplate reader was developed. This method is 8 times more effective than basic technique in one cycle per person. Futhermore, considering for less than 1/10 volume of materials required for ALS test and enzyme kinetics with 16 times faster speed compared to those of former procedure, this HTS method has more than 100 times higher efficacy than basic system in a consecutive procedure. We discovered 11 new ALS inhibitors such as 2-oxoglutaric acid, aminooxyacetic acid, azelaic acid, citric acid, cyanuric fluoride, itaconic acid, malonic acid, niclosamide, oxalic acid, glyoxylic acid, and suramin from 107 commercial plant-specific inhibitors using this technique. We hope these results might be useful to discover lead compounds for developing new novel herbicide candidate.

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Comparison of Pap Smear Screening Results between Akha Hill Tribe and Urban Women in Chiang Rai Province, Thailand

  • Kritpetcharat, Onanong;Wutichouy, Wiwat;Sirijaichingkul, Suchat;Kritpetcharat, Panutas
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.11
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    • pp.5501-5504
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    • 2012
  • Cervical cancer is an important woman's health problems worldwide, especially in low socio-economic countries. The aim of this study was to compare the Pap smear screening results between Akha hill tribe and urban women who live in Chiang Rai province, Thailand. Screening was conducted for 1,100 Akha women and 1,100 urban women who came to have the Pap smear at Chiangrai Prachanukroh Hospital and 1 private cytology laboratory from January to June 2008. The demographic characteristics and factors related to abnormal Pap smears of these women were gathered using closed model questionnaires. Abnormal Pap smears were defined according to the Bethesda 2001 system. The results showed that the prevalence of abnormal Pap smears was 12.2% in Akha women and 4.5% in urban women. The highest prevalence of Pap abnormalities was found in the 41-50 years age group in both populations (4.5% in Akha and 1.7% in urban women). In both populations, abnormal Pap smears were found in <21 years age groups. From the questionnaires, the possible risk factors related to the higher prevalence of abnormal Pap smears in Akha women were early age at marriage (${\leq}17$ years), high frequency pregnacies and high parity and no/low education level. In conclusion, cervical cancer control by education and early detection by Pap smear screening is necessary for hill tribe women. More Pap smear screening service units should be set to improve the coverage for the risk group women who got married in young age, especial in ethnic groups.

Development of a highly effective T-DNA inserted mutant screening method in a Chinese cabbage (Brassica rapa L. spp. pekinensis) reverse genetics system

  • Lee, Gi-Ho;Kang, Yoon-Jee;Yi, Seul-Ki;Lim, Suk-Bin;Park, Young-Doo
    • Plant Biotechnology Reports
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    • v.4 no.3
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    • pp.201-211
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    • 2010
  • We present a highly effective T-DNA inserted gene screening method as part of a reverse genetics model system using the Chinese cabbage (Brassica rapa L. spp. pekinensis). Three-step two-dimensional (2D) matrix strategies are potentially accurate and useful for the identification of specific T-DNA inserted mutants from a large population. To construct our Chinese cabbage model, we utilized a forward genetics screening approach for the abnormal phenotypes that were obtained from transgenic plants of Brassica rapa generated with Agrobacteria tumefaciens containing the pRCV2 vector. From one transgenic plant with an abnormal phenotype, we observed that the st1 gene (which is related to senescence-associated process proteins) contained a T-DNA fragment, and that its expression level was decreased. This T-DNA insert was then used as a control to construct an effective screening pool. As a result, the optimum template concentration was found to be 0.1-1 ng in our PCR strategy. For other conditions, positive changes to the Gibbs free energy prevented the formation of oligo dimers and hairpin loop structures, and autosegment extension gave better results for long fragment amplification. Using this effective reverse genetics screening method, only 23 PCR reactions were necessary to select a target gene from a pool of 100 individual DNAs. Finally, we also confirmed that the sequence we obtained from the above method was identical to the flanking sequence isolated by rescue cloning.

Screening of Anti-Adhesion Agents for Pathogenic Escherichia coli O157:H7 by Targeting the GrlA Activator

  • Sin Young Hong;Byoung Sik Kim
    • Journal of Microbiology and Biotechnology
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    • v.33 no.3
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    • pp.329-338
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    • 2023
  • Enterohemorrhagic Escherichia coli (EHEC) is a foodborne pathogen that produces attaching and effacing lesions on the large intestine and causes hemorrhagic colitis. It is primarily transmitted through the consumption of contaminated meat or fresh produce. Similar to other bacterial pathogens, antibiotic resistance is of concern for EHEC. Furthermore, since the production of Shiga toxin by this pathogen is enhanced after antibiotic treatment, alternative agents that control EHEC are necessary. This study aimed to discover alternative treatments that target virulence factors and reduce EHEC toxicity. The locus of enterocyte effacement (LEE) is essential for EHEC attachment to host cells and virulence, and most of the LEE genes are positively regulated by the transcriptional regulator, Ler. GrlA protein, a transcriptional activator of ler, is thus a potential target for virulence inhibitors of EHEC. To identify the GrlA inhibitors, an in vivo high-throughput screening (HTS) system consisting of a GrlA-expressing plasmid and a reporter plasmid was constructed. Since the reporter luminescence gene was fused to the ler promoter, the bioluminescence would decrease if inhibitors affected the GrlA. By screening 8,201 compounds from the Korea Chemical Bank, we identified a novel GrlA inhibitor named Grlactin [3-[(2,4-dichlorophenoxy)methyl]-4-(3-methylbut-2-en-1-yl)-4,5-dihydro-1,2,4-oxadiazol-5-one], which suppresses the expression of LEE genes. Grlactin significantly diminished the adhesion of EHEC strain EDL933 to human epithelial cells without inhibiting bacterial growth. These findings suggest that the developed screening system was effective at identifying GrlA inhibitors, and Grlactin has potential for use as a novel anti-adhesion agent for EHEC while reducing the incidence of resistance.

Aequorin Based Functional Assessment of the Melanin Concentrating Hormone Receptor by Intracellular Calcium Mobilization

  • Lee, Sung-Hou
    • Biomolecules & Therapeutics
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    • v.18 no.2
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    • pp.152-158
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    • 2010
  • Melanin concentrating hormone is a neuropeptide highly expressed in the brain that regulates several physiological functions mediated by receptors in the G-protein coupled receptor family, especially plays an important role in the complex regulation of energy balance and body weight mediated by the melanin concentrating hormone receptor subtype 1 (MCH1). Compelling pharmacological evidence implicating MCH1 signaling in the regulation of food intake and energy expenditure has generated a great deal of interest by pharmaceutical companies as MCH1 antagonists may have potential therapeutic benefit in the treatment of obesity and metabolic syndrome. Although fluorescence-based calcium mobilization assay platform has been one of the most widely accepted tools for receptor research and drug discovery, fluorescence interference and shallow assay window limit their application in high throughput screening and have led to a growing interest in alternative, luminescence-based technologies. Herein, a luminescence-based functional assay system for the MCH1 receptor was developed and validated with the mitochondrial targeted aequorin. Aequorin based functional assay system for MCH1 presented excellent Z' factor (0.8983) and high signal-to-noise ratio (141.9). The nonpeptide MCH1 receptor antagonist, SNAP 7941 and GSK 803430, exhibited $IC_{50}$ values of 0.62 ${\pm}$ 0.11 and 12.29 ${\pm}$ 2.31 nM with excellent correlation coefficient. These results suggest that the aequorin based assay system for MCH1 is a strong alternative to the traditional GPCR related tools such as radioligand binding experiments and fluorescence functional determinations for the compound screening and receptor research.

Development of Digital Fault Detection Systems for Screening Open and Short of Wire Harness (와이어 하네스 단선 단락 선별을 위한 디지털 고장 검출 시스템 개발)

  • A Ran Kim;Jae Wan Park;Ha Seon Kim;Jae Hoon Jeong;Sun Young Kim
    • Journal of Drive and Control
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    • v.20 no.4
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    • pp.140-149
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    • 2023
  • Wire harness is a component for efficient control when electronic parts are required such as construction machinery and electric vehicles. With emerging issues such as autonomous driving and automation in construction, a wire harness composed of multiple cables has become an essential part because more electronic parts are required. However, when a wire harness failure occurs, systems can be stopped, accidents can occur, and economic damage can be significant. Therefore, in this paper, we developed a digital fault screening system that could easily and quickly diagnose faults in the wire harness. The principle of the developed system was to sequentially send pulse signals to the wire harness and use returned signals to perform fault detection. As a result of diagnosing faults using the developed failure detection system, a detection accuracy of 99.9 % was confirmed through the experiments.

Factors Related to Breast Cancer Screening using Mammography and Clinical Breast Examination of Korean Women Over 40 Years of Age (한국 도시여성의 유방촬영술과 유방임상검사 관련요인)

  • Eun, Young;Gu, Mee-Ok;Lee, Eunice L.;Choi, Kyung-Sook;Jun, Myung-Hee
    • Korean Journal of Adult Nursing
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    • v.21 no.6
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    • pp.580-592
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    • 2009
  • Purpose: The purpose of this study was to investigate the factors related to Breast cancer Screening using Mammography and CBE of Korean women over 40 years of age. Methods: The participants for this study were 183 Korean women living in 3 urban cities and aged from 40 to 75. The data were collected using structured questionnaires which included sociodemographic factors (11 items), frequency and regularity of mammography and clinical breast examination (7 items), knowledge (16 items), health belief model scale (28 items), and family support (4 items) about breast cancer and breast cancer screening. Frequencies, Chronbach's alpha for reliability, Chi-square, t-test and logistic regression with the SPSS/WIN 12 program were used to analyze the data. Results: The percentages of Korean women who had a mammography and CBE for breast cancer screening were 60.1 and 31.1, respectively. Logistic regression analyses demonstrated that regular check ups and perceived barriers were significant predictors of mammography and CBE use for breast cancer screening. Conclusion: In order to increase the frequency of breast cancer screening practices, educational support and a health care delivery system is needed to improve the chance of regular health check ups.

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Cervical Cancer Screening: Recommendations for Muslim Societies

  • Khodakarami, Nahid;Farzaneh, Farah;Yavari, Parvin;Akbari, Mohamad Esmaeil
    • Asian Pacific Journal of Cancer Prevention
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    • v.17 no.1
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    • pp.239-247
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    • 2016
  • Background: The overall cervical cancer incidence rate is low in Iran; however, because of a higher risk of death for cervical cancer, a disease that kills women in middle age, a cervical cancer control program is needed. The aim of this study was to provide consensus recommendation for cervical cancer prevention in Iran and other Muslim societies with low incidences of cervical cancer. Materials and Methods: Through a practical guideline development cycle, we developed six questions that were relevant to produce the recommendation. We reviewed 190 full text records of cervical cancer in Iran (1971 to 2013) of which 13 articles were related to the data needed to answer the recommendation questions. We also reviewed World Health Organization, IARC, GLOBOCAN report, Iran Ministry of Health cancer registry report and 8 available foreign countries guidelines. Lastly, we also evaluated the Pap smear results of 825 women who participated in the Iranian HPV survey, of whom 328 were followed-up after a 5-year interval. Results: The obtained data highlighted the burden of HPV and cervical cancer situation in Iran. Experts emphasized the necessity of a cervical cancer screening program for Iranian women, and recommended an organized screening program with a cytological evaluation (Pap smear) that would start at the age of 30 years, repeated every 5 years, and end at the age of 69 years. Meanwhile, there is no need for screening among women with a hysterectomy, and screening should be postponed to post-partum among pregnant women. Conclusions: An organized cervical cancer screening is a necessity for Iran as more than 500-900 women in middle age diagnosed with an invasive cervical cancer every year cannot be ignored. This recommendation should be taken into account by the National Health System of Iran and Muslim countries with shared culture and behavior patterns. CUBA HPV test could be consideration in countries Muslim country with appropriate budget, resources and facility.

The Development and Effect of Navigator Education Program for Cancer Screening on Women in the Community (지역사회 여성암 검진 네비게이터 교육 프로그램 개발 및 효과 분석)

  • Lee, Bo-Young;Jo, Heui-Sug;Lee, Hey-Jean
    • Journal of agricultural medicine and community health
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    • v.34 no.2
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    • pp.214-222
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    • 2009
  • Objectives: This study was performed to evaluate the effect of navigator education program for cancer screening, which is designed for improvement in knowledge of cancer, perceived self efficacy and communication skill of the breast and cervical cancer screening for middle-aged and aged women in urban areas. Cancer screening navigator is lay health advisor who are educated for providing information, emotional support about cancer screening at the community. Methods: The subjects were 33 women at the age of 40-69 and educated for 12 hours through the education program. The control group subjects were 30 women. For statistical analysis, descriptive statistics and paired t-test were used with SPSS WIN 14.0. Results: Contents of education program were case of cancer early detection, benefit of breast cancer screening, benefit of cervical cancer screening, health care system for cancer screening, role of cancer screening navigator, communication skill, transtheoretical model and role play. Knowledge of cancer(t=4.267, p=0.000) and communication skill(t=4.947, p=0.000) of the women increased significantly after implementing the 12 hours education program. Conclusion: The results suggest that navigator education for cancer screening has an effect in increasing knowledge of cancer, and communication skill scores.