• Journal of Microbiology and Biotechnology
• /
• v.20 no.11
• /
• pp.1563-1570
• /
• 2010

To construct a highly sensitive detection system for endocrine disruptors (EDs), we have compared the activity of promoters with the n-alkane-inducible cytochrome P450 gene (ALK1), isocitrate lyase gene (ICL1), ribosomal protein S7 gene (RPS7), and the translation elongation factor-1${\alpha}$ gene (TEF1) for the heterologous gene in Yarrowia lipolytica. The promoters were introduced into the upstream of the lacZ or hERa reporter genes, respectively, and the activity was evaluated by ${\beta}$-galactosidase assay for lacZ and Western blot analysis for hER${\alpha}$. The expression analysis revealed that the ALK1 and ICL1 promoters were induced by n-decane and by EtOH, respectively. The constitutive promoter of RPS7 and TEF1 showed mostly a high level of expression in the presence of glucose and glycerol, respectively. In particular, the TEF1 promoter showed the highest ${\beta}$-galactosidase activity and a significant signal by Western blotting with the anti-estrogen receptor, compared with the other promoters. Moreover, the detection system was constructed with promoters linked to the upstream of the expression vector for the hER${\alpha}$ gene transformed into the Y. lipolytica with a chromosome-integrated lacZ reporter gene under the control of estrogen response elements (EREs). It was indicated that a combination of pTEF1p-hER${\alpha}$ and CXAU1-2XERE was the most effective system for the $E_2$-dependent induction of the ${\beta}$-galactosidase activity. This system showed the highest ${\beta}$-galactosidase activity at $10^{-6}\;M\;E_2$, and the activity could be detected at even the concentration of $10^{-10}\;M\;E_2$. As a result, we have constructed a strongly sensitive detection system with Y. lipolitica to evaluate recognized/suspected ED chemicals, such as natural/synthetic hormones, pesticides, and commercial chemicals. The results demonstrate the utility, sensitivity, and reproducibility of the system for identifying and characterizing environmental estrogens.

• Annual Conference of KIPS
• /
• 2007.05a
• /
• pp.1487-1490
• /
• 2007

디지털 위성방송의 시작과 더불어 본격적인 데이터 방송의 시대가 열렸다. 데이터방송이 시작 되면서 데이터방송용 양방향 콘텐츠에 대한 수요가 급속하게 증가하고 있다. 하지만 양방향 콘텐츠 개발에 필요한 저작 도구 및 검증 시스템은 아주 초보적인 수준에 머물러 있는 것이 현실이다. 그러나 방송의 특성상 콘텐츠 상에서의 오류는 방송 사고에까지 이를 수 있는 심각한 상황이 연출 될 수 있다. 본 연구 팀은 이러한 DTV 콘텐츠 개발 요구에 부응하여, 개발자의 콘텐츠 개발 및 사업자 또는 기관에서의 콘텐츠 검증이 원활이 이루어 질수 있도록 하는 양방향 콘텐츠 검증 시스템을 개발 중이다. 양방향 콘텐츠 검증 시스템은 Java 컴파일러, 디버거, 미들웨어, 가상머신, 그리고 IDE 등으로 구성된다. 본 논문에서 제시한 자바 컴파일러는 양방향 콘텐츠 검증 시스템에서 데이터 방송용 자바 애플리케이션(Xlet)을 컴파일하여 에뮬레이팅 하거나 런타임 상에서 디버깅이 가능하도록 하는 바이너리형태의 class 파일을 생성한다. 이를 위해 Java 컴파일러는 *.java 파일을 입력으로 받아 어휘 분석과 구문 분석 과정을 거친 후 SDT(syntax-directed translation)에 의해 AST(Abstract Syntax Tree)를 생성한다. 클래스링커는 생성된 AST를 탐색하여 동적으로 로딩 되는 파일들을 연결하여 AST를 확장한다. 의미 분석과정에서는 확장된 AST를 입력으로 받아 참조된 명칭의 사용이 타당한지 등을 검사하고 코드 생성이 용이하도록 AST를 변형하고 부가적인 정보를 삽입하여 ST(Semantic Tree)를 생성한다. 코드 생성 단계에서는 ST를 입력으로 받아 이미 정해 놓은 패턴에 맞추어 Bytecode를 출력한다.ovoids에서도 각각의 점들에 대한 선량을 측정하였다. SAS와 SSAS의 직장에 미치는 선량차이는 실제 임상에서의 관심 점들과 가장 가까운 25 mm(R2)와 30 mm(R3)거리에서 각각 8.0% 6.0%였고 SAS와 FWAS의 직장에 미치는 선량차이는 25 mm(R2) 와 30 mm(R3)거리에서 각각 25.0% 23.0%로 나타났다. SAS와 SSAS의 방광에 미치는 선량차이는 20 m(Bl)와 30 mm(B2)거리에서 각각 8.0% 3.0%였고 SAS와 FWAS의 방광에 미치는 선량차이는 20 mm(Bl)와 30 mm(B2)거리에서 각각 23.0%, 17.0%로 나타났다. SAS를 SSAS나 FWAS로 대체하였을 때 직장에 미치는 선량은 SSAS는 최대 8.0 %, FWAS는 최대 26.0 %까지 감소되고 방광에 미치는 선량은 SSAS는 최대 8.0 % FWAS는 최대 23.0%까지 감소됨을 알 수 있었고 FWAS가 SSAS 보다 차폐효과가 더 좋은 것으로 나타났으며 이 두 종류의 shielded applicator set는 부인암의 근접치료시 직장과 방광으로 가는 선량을 감소시켜 환자치료의 최적화를 이룰 수 있을 것으로 생각된다.)한 항균(抗菌) 효과(效果)를 나타내었다. 이상(以上)의 결과(結果)로 보아 선방활명음(仙方活命飮)의 항균(抗菌) 효능(效能)은 군약(君藥)인 대황(大黃)의 성분(成分) 중(中)의 하나인 stilbene 계열(系列)의 화합물(化合物)인 Rhapontigenin과 Rhaponticin의 작용(作用)에 의(依)한 것이며, 이는 한의학(韓醫學) 방제(方劑) 원리(原理)인 군신좌사(君臣佐使) 이론(理論)에서 군약(君藥)이 주증(主症)에 주(主)로 작용(作用)하는 약물(藥物)이라는 것을 밝혀주는 것이라고

• Journal of Sericultural and Entomological Science
• /
• v.37 no.1
• /
• pp.46-51
• /
• 1995

In order to develope baculovirus expression vector system, we constructed new transfer vector of nuclear polyhedrosis virus of the silkworm, Bombyx mori. The promoter region containing only adenine of translation start codon of polyhedrin gene was cloned by polymerase chain reaction technique. And the 5' and 3' leader regions of polyhedrin gene was sequentially cloned. The polyhedrin coding gene was deleted from the ＋2 to the ＋597 position. As the result, we constructed new transfer vector which has EcoRI, SacI and KpnI sites for the cloning sites of foreign gene. New transfer vector was named as pBmKSKl. Escherichia coli $\beta$-galactosidase gene as foreign gene was inserted into pBmKSKl, under the control of the polyhedrin promoter and expressed in B. mori cells. The result showed that the new transfer vector pBmKSK1 is functional.

• Research in Plant Disease
• /
• v.22 no.2
• /
• pp.111-115
• /
• 2016

Wilt disease appeared the first in greenhouse-grown safflower (Carthamus tinctorius) in Jeonju, Korea. With the advancement of the disease, the infected plants were withered and died. In order to investigate the causal organism of this symptom disease, fungus was isolated from the infected plants and cultured on potato dextrose agar medium. The fungus showed the white or orange colony color with aerial mycelium. Macroconidia were from falcate to straight, usually 3-5 septate with $38.0-66.7{\times}2.9-4.4{\mu}m$. The fungus was inoculated to a new safflower plant and caused the same wilt. With morphological characters and pathogenicity results, sequence analyses (internal transcribed spacer ribosomal DNA and translation elongation factor $1{\alpha}$) suggested that, the isolated fungus is Fusarium proliferatum. This is the first report of Fusarium wilt disease caused by F. proliferatum on safflower in Korea.

• Journal of Korean Foot and Ankle Society
• /
• v.21 no.1
• /
• pp.12-16
• /
• 2017

Purpose: Anterior drawer and varus stress radiographs are commonly to diagnose chronic lateral ankle instability. We compared the preoperative stress radiographs with the intraoperative radiographs under anesthesia to determine the accuracy and efficacy of stress radiographs in an outpatient clinical environment. Materials and Methods: Data was collected from patients who underwent a modified $Brostr{\ddot{o}}m$ operation for painful chronic unilateral lateral ankle instability between January 2014 and June 2016. Subjects were divided into three groups-complete tear, partial tear, and instability without rupture-according to the status of preoperative MRI findings of the anterior talofibular ligament. The anterior drawer and varus stress radiographs were taken preoperatively and intraoperatively under anesthesia. Results: Ninety-six patients, with a mean age of 29.63 years, were enrolled. There were 39, 46, and 11 patients in the complete tear, partial tear, and instability without rupture groups, respectively. On the anterior drawer and varus stress radiographs of the affected limb, talar anterior translation and varus tilting were significantly increased by 2.56 mm and $2.0^{\circ}$. The gaps between the unaffected limbs were also increased by 2.47 mm and $1.32^{\circ}$ after anesthesia. Although the stress radiographs were taken under anesthesia, the results were often smaller than the diagnostic value. Conclusion: Stress radiographs for painful chronic lateral ankle instability taken at the outpatient clinic might be inaccurate for diagnosis.

• Journal of Microbiology and Biotechnology
• /
• v.8 no.6
• /
• pp.676-684
• /
• 1998

Baculovirus transfer and expression vectors with Hyphantria cunea nuclear polyhedrosis virus (HcNPV) were constructed. An initial transfer vector, pHcEV, constructed using HcNPV was previously reported (Park et al. 1993. J. Kor. Soc. Viral. 23: 141-151). Herein, the size of the vector was properly reduced, and a functionally perfect vector was constructed and named pHcEV-IV (6.7 kb). The vector has a 2.2-kb HcNPV DNA sequence in the 5'-flanking region of the vector's polyhedrin gene promoter. The 1.8-kb HcNPV DNA sequence, poly A signal sequence, T3 primer sequence, and 13 multicloning site sequences, in order, were ligated in front of the translation start codon of the polyhedrin gene. The cloning indicating marker lacZ gene was inserted into the pHcEV-IV, named pHcEV-IV-lacZ, and transferred into the wild-type virus. Recombinant expression virus, lacZ-HcNPV, was constructed by replacing the lacZ gene in the pHcEV-IV-lacZ with the polyhedrin gene of the wild-type virus. The recombinant virus was isolated from blue plaques that produce $\beta$-galactosidase without polyhedra. The lacZ gene insertion was confirmed by Southern hybridization analysis. The expression of the lacZ gene in Spodoptera frugiperda cells infected with the lacZ-HcNPV was examined by SDS-PAGE and colorimetric assay. One 116-kDa LacZ protein band appeared on the PAGE. The production rate of the $\beta$-galactosidase was approximately 50 international units (IU) per min per ml between 2 to 5 days postinfection (p.i.). The highest activity occurred at five days p.i. was 170 IU/min/$m\ell$. The enzyme activity first appeared about 20 h p.i. as measured by colorimetric assay.

• Molecules and Cells
• /
• v.45 no.9
• /
• pp.660-672
• /
• 2022

The target of rapamycin complex (TORC) plays a key role in plant cell growth and survival by regulating the gene expression and metabolism according to environmental information. TORC activates transcription, mRNA translation, and anabolic processes under favorable conditions, thereby promoting plant growth and development. Tomato fruit ripening is a complex developmental process promoted by ethylene and specific transcription factors. TORC is known to modulate leaf senescence in tomato. In this study, we investigated the function of TORC in tomato fruit ripening using virus-induced gene silencing (VIGS) of the TORC genes, TOR, lethal with SEC13 protein 8 (LST8), and regulatory-associated protein of TOR (RAPTOR). Quantitative reverse transcription-polymerase chain reaction showed that the expression levels of tomato TORC genes were the highest in the orange stage during fruit development in Micro-Tom tomato. VIGS of these TORC genes using stage 2 tomato accelerated fruit ripening with premature orange/red coloring and decreased fruit growth, when control tobacco rattle virus 2 (TRV2)-myc fruits reached the mature green stage. TORC-deficient fruits showed early accumulation of carotenoid lycopene and reduced cellulose deposition in pericarp cell walls. The early ripening fruits had higher levels of transcripts related to fruit ripening transcription factors, ethylene biosynthesis, carotenoid synthesis, and cell wall modification. Finally, the early ripening phenotype in Micro-Tom tomato was reproduced in the commercial cultivar Moneymaker tomato by VIGS of the TORC genes. Collectively, these results demonstrate that TORC plays an important role in tomato fruit ripening by modulating the transcription of various ripening-related genes.

• Journal of Plant Biotechnology
• /
• v.37 no.1
• /
• pp.89-95
• /
• 2010

Dihydroflavonol 4-reductase (DFR) is a key enzyme of the flavonoid biosynthesis pathway which catalyses the NADPH-dependent reduction of 2R,3R-trans-dihydroflavonols to leucoanthocyanidins. In this study we describe cloning and expression of the genes encoding the flavonoid-biosynthetic enzyme DFR in Gypsophila paniculata L. Inspection of the 1279 bp long sequence revealed an open reading frame 1063 bp, including a 36 bp 5' leader region and 181 bp 3' untranslated region. Comparison of the coding region of this DFR cDNA sequence including the sequences of Arabidopsis thaliana, Citrus sinensis, Dianthus caryophyllus, Ipomoea batatas, Matthiola incana, Nierembergia sp, Petunia hybrida, Solanum tuberosum, Vitis vinifera reveals an identity higher than 69% at the nucleotide level. The function of this nucleotide sequences was verified by comparison with amino acid sequences of the amino-terminus and tryptic peptides from purified plant enzyme, by northern blotting with mRNA from wild type and mutant plants, by in vitro expression yielding and enzymatically active reductase, as indicated by the small leucopelargonidin peak. Genomic southern blot analysis showed the presence of only one gene for DFR in Gypsophila paniculata.

• The Korean Journal of Physiology and Pharmacology
• /
• v.10 no.5
• /
• pp.263-271
• /
• 2006

Since astrocytes were shown to play a central role in maintaining neuronal viability both under normal conditions and during stress such as ischemia, studies of the astrocytic response to stress are essential to understand many types of brain pathology. The micro array system permitted screening of large numbers of genes in biological or pathological processes. Therefore, the gene expression patterns in the in vitro model of astrocytes following exposure to oxygen-glucose deprivation (OGD) were evaluated by using the micro array analysis. Primary astrocytic cultures were prepared from postnatal Swiss Webster mice. The cells were exposed to OGD for 4 hrs at $37^{\circ}C$ prior to cell harvesting. From the cultured cells, we isolated mRNA, synthesized cDNA, converted to biotinylated cRNA and then reacted with GeneChips. The data were normalized and analyzed using dChip and GenMAPP tools. After 4 hrs exposure to OGD, 4 genes were increased more than 2 folds and 51 genes were decreased more than 2 folds compared with the control condition. The data suggest that the OGD has general suppressive effect on the gene expression with the exception of some genes which are related with ischemic cell death directly or indirectly. These genes are mainly involved in apoptotic and protein translation pathways and gap junction component. These results suggest that microarray analysis of gene expression may be useful for screening novel molecular mediators of astrocyte response to ischemic injury and making profound understanding of the cellular mechanisms as a whole. Such a screening technique should provide insights into the molecular basis of brain disorders and help to identify potential targets for therapy.

• Molecules and Cells
• /
• v.43 no.9
• /
• pp.821-830
• /
• 2020

Altered dendritic morphology is frequently observed in various neurological disorders including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), but the cellular and molecular basis underlying these pathogenic dendritic abnormalities remains largely unclear. In this study, we investigated dendritic morphological defects caused by dipeptide repeat protein (DPR) toxicity associated with G4C2 expansion mutation of C9orf72 (the leading genetic cause of ALS and FTD) in Drosophila neurons and characterized the underlying pathogenic mechanisms. Among the five DPRs produced by repeat-associated non-ATG translation of G4C2 repeats, we found that arginine-rich DPRs (PR and GR) led to the most significant reduction in dendritic branches and plasma membrane (PM) supply in Class IV dendritic arborization (C4 da) neurons. Furthermore, expression of PR and GR reduced the number of Golgi outposts (GOPs) in dendrites. In Drosophila brains, expression of PR, but not GR, led to a significant reduction in the mRNA level of CrebA, a transcription factor regulating the formation of GOPs. Overexpressing CrebA in PR-expressing C4 da neurons mitigated PM supply defects and restored the number of GOPs, but the number of dendritic branches remained unchanged, suggesting that other molecules besides CrebA may be involved in dendritic branching. Taken together, our results provide valuable insight into the understanding of dendritic pathology associated with C9-ALS/FTD.