• Title/Summary/Keyword: library screening

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Ttrosine Hydroxylase in Japanese Medaka (Oryzias latipes): cDNA Cloning and Molecular Monitoring of TH Gene Expression As a Biomarker (송사리 Tyrosine Hydroxylase: cDNA 클로닝 및 생물지표로서의 TH 유전자 발현의 분자생물학적 추적)

  • Shin, Sung-Woo;Kim, Jung-Sang;Chon, Tae-Soo;Lee, Sung-Kyu;Koh, Sung-Cheol
    • Environmental Analysis Health and Toxicology
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    • v.15 no.4
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    • pp.131-137
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    • 2000
  • The release of hazardous waste materials into the environment poses serious risks in humans and ecosystems. The risk assessment of environmental pollutants including hazardous chemicals requires a comprehensive measurement of hazard and exposure of the chemicals that can be achieved by toxicity evaluation using a biological system such as biomarkers. In this report we have tried to develop a biomarker used to elucidate a molecular basis of, and to monitor abnormal behaviors caused by diazinon in Japanese medaka (Oryzias latipes) as a model organism. First, an attempt was made to clone tyrosine hydroxylase gene from Japanese medaka that would be a candidate for a biomarker for neuronal modulations and behaviors. For monitoring experiments at behavioral and molecular biological levels, the fish were treated under different sublethal conditions of diazinon and their behavioral responses were observed . In this study we have successfully cloned a partial TH gene from the medaka fish through PCR screening of an ovary cDNA library. DNA sequencing analysis revealed that the amplified fragment was 327 bp encoding 109 amino acids. Comparing the DNA sequence of medaka TH with other species, TH gene revealed the DNA sequence was completely identical to that of rat TH. In the RT-PCR, 330 Up of mRNA was consistently amplified in all the treated samples including control There were no significant differences in the TH expression level regardless of treating concentrations (1∼5,000 ppb) and time (0∼48 hr) The reason appeared to be that RT-PCR was not performed using through a quantitative analysis normalized against an actin gene expression. Organ or tissue - specific detection of TH activity and mRNA as biomarkers will be a useful monitoring tool for neurobehavioral changes in fish influenced by toxic chemicals. Furthermore, quantitative analysis of locomotive patterns and its correlation with the neurochemical and molecular data would be highly useful in measuring toxicity and hazard ofvarious environmental pollutants.

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Cloning and Transcription Analysis of Sporulation Gene (spo5) in Schizosaccharomyces pombe (Schizosaccharomyces bombe 포자형성 유전자(spo5)의 Cloning 및 전사조절)

  • 김동주
    • The Korean Journal of Food And Nutrition
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    • v.15 no.2
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    • pp.112-118
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    • 2002
  • Sporulation in the fission yeast Schizosaccharomyces pombe has been regarded as an important model of cellular development and differentiation. S. pombe cells proliferate by mitosis and binary fission on growth medium. Deprivation of nutrients especially nitrogen sources, causes the cessation of mitosis and initiates sexual reproduction by matting between two sexually compatible cell types. Meiosis is then followed in a diploid cell in the absence of nitrogen source. DNA fragment complemented with the mutations of sporulation gene was isolated from the S. pombe gene library constructed in the vector, pDB 248' and designated as pDB(spo5)1. We futher analyzed six recombinant plasmids, pDB(spo5)2, pDB(spo5)3, pDB(spo5)4, pDB(spo5)5, pDB (spo5)6, pDB(spo5)7 and found each of these plasmids is able to rescue the spo5-2, spo5-3, spo5-4, spo5-5, spo5-6, spo5-7 mutations, respectively. Mapping of the integrated plasmid into the homologous site of the S. pombe chromosomes demonstrated that pDB(spo5)1, and pDB(spu5)Rl contained the spo5 gene. Transcripts of spo5 gene were analyzed by Northern hybridization. Two transcripts of 3.2 kb and 2.5kb were detected with 5kb Hind Ⅲ fragment containing a part of the spo5 gene as a probe. The small mRNA(2.5kb) appeared only when a wild-type strain was cultured in the absence of nitrogen source in which condition the large mRNA (3.2kb) was produced constitutively. Appearance of a 2.5kb spo5-mRNA depends upon the function of the meil, mei2 and mei3 genes.

Mechanisms of Insulinotropic Effect of YHB-2017 [Genistein] Isolated from fermentation Broths of Streptomyces sp. (방선균에서 유래한 YHB-2017 [Genistein]의 인슐린 분비 촉진 작용 기전)

  • Kwag, Won-Jae;Park, You-Hoi;Park, Jun-Chul;Lee, Byung-Kyu;Kang, Yup;Choe, Tae-Boo
    • KSBB Journal
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    • v.21 no.6 s.101
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    • pp.466-473
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    • 2006
  • Impaired insulin secretion from pancreatic beta-cells in response to glucose is an important feature in the pathology of non-insulin-dependent diabetes mellitus (NIDDM). In the course of screening for useful insulin secretagogues, we have isolated and identified YHB-2017 (Genistein) as a insulin secretion potentiator from fermentation broths of our in-house microbial library. The insulinotropic activity of YHB-2017 in isolated rat pancreatic islets was exerted only at high concentration of glucose (8.3-16 mM) but not at low concentration of glucose (3.3-5.5 mM). Also, in perifusion study with isolated rat pancreatic islets, YHB-2017 stimulated insulin secretion in a time-dependent manner when YHB-2017 was added to KRB buffer containing 16 mM glucose. In the presence of $200\;{\mu}M$ diazoxide and 35 mM KCI, which stimulates maximum $Ca^{2+}$ influx independently of KATP channel, YHB-2017 enhanced KATP channel-independent insulin secretion at high concentration glucose (16 mM). To elucidate the mechanisms of the glucose-dependent potentiation effect of YHB-2017, pharmacologic inhibitors for protein kinase A, protein kinase C and calcium/calmodulin kinase II were pre-treated and then the potentiation effect of YHB-2017 on insulin secretion was investigated. Pre-treatment of H89 as a PKA inhibitor had a significant inhibitory effect on YHB-2017-induced potentiation effect. Furthermore, western immunoblotting analyses revealed that YHB-2017 increased phosphorylation of PKA substrates and cAMP response element-binding protein (CREB) under high concentration of glucose. These results demonstrated that the insulinotropic effect of YHB-2017 is mediated through PKA signal pathway and activated amplifying $K_{ATP}$ channel-independent insulin secretion pathway.

Korean Practice Guidelines for Gastric Cancer 2022: An Evidence-based, Multidisciplinary Approach

  • Tae-Han Kim;In-Ho Kim;Seung Joo Kang;Miyoung Choi;Baek-Hui Kim;Bang Wool Eom;Bum Jun Kim;Byung-Hoon Min;Chang In Choi;Cheol Min Shin;Chung Hyun Tae;Chung sik Gong;Dong Jin Kim;Arthur Eung-Hyuck Cho;Eun Jeong Gong;Geum Jong Song;Hyeon-Su Im;Hye Seong Ahn;Hyun Lim;Hyung-Don Kim;Jae-Joon Kim;Jeong Il Yu;Jeong Won Lee;Ji Yeon Park;Jwa Hoon Kim;Kyoung Doo Song;Minkyu Jung;Mi Ran Jung;Sang-Yong Son;Shin-Hoo Park;Soo Jin Kim;Sung Hak Lee;Tae-Yong Kim;Woo Kyun Bae;Woong Sub Koom;Yeseob Jee;Yoo Min Kim;Yoonjin Kwak;Young Suk Park;Hye Sook Han;Su Youn Nam;Seong-Ho Kong;The Development Working Group for the Korean Practice Guidelines for Gastric Cancer 2022 Task Force Team
    • Journal of Gastric Cancer
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    • v.23 no.1
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    • pp.3-106
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    • 2023
  • Gastric cancer is one of the most common cancers in Korea and the world. Since 2004, this is the 4th gastric cancer guideline published in Korea which is the revised version of previous evidence-based approach in 2018. Current guideline is a collaborative work of the interdisciplinary working group including experts in the field of gastric surgery, gastroenterology, endoscopy, medical oncology, abdominal radiology, pathology, nuclear medicine, radiation oncology and guideline development methodology. Total of 33 key questions were updated or proposed after a collaborative review by the working group and 40 statements were developed according to the systematic review using the MEDLINE, Embase, Cochrane Library and KoreaMed database. The level of evidence and the grading of recommendations were categorized according to the Grading of Recommendations, Assessment, Development and Evaluation proposition. Evidence level, benefit, harm, and clinical applicability was considered as the significant factors for recommendation. The working group reviewed recommendations and discussed for consensus. In the earlier part, general consideration discusses screening, diagnosis and staging of endoscopy, pathology, radiology, and nuclear medicine. Flowchart is depicted with statements which is supported by meta-analysis and references. Since clinical trial and systematic review was not suitable for postoperative oncologic and nutritional follow-up, working group agreed to conduct a nationwide survey investigating the clinical practice of all tertiary or general hospitals in Korea. The purpose of this survey was to provide baseline information on follow up. Herein we present a multidisciplinary-evidence based gastric cancer guideline.