• Molecules and Cells
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• 제21권1호
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• pp.21-28
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• 2006

The interferon-induced, double-stranded RNA (dsRNA)-dependent protein kinase PKR plays a key role in interferon-mediated host defense against viral infection, and is implicated in cellular transformation and apoptosis. We have isolated a cDNA of simian PKR encoding a product with 83% amino acid identity to the human homolog and showed that PKR expression is significantly attenuated in the Vero E6 African green monkey kidney cells devoid of type I interferon genes. A variant form of PKR lacking the exon 12 in the kinase domain is produced in these cells, presumably from an alternatively spliced transcript. Unlike wild type PKR, the variant protein named PKR-${\Delta}E12$ is incapable of auto-phosphorylation and phosphorylation of eIF2-${\alpha}$, indicating that the kinase sub-domains III and IV embedded in exon 12 are indispensable for catalytic function. PKR-${\Delta}E12$ had no dominant negative effect but was weakly phosphorylated in trans by wild type PKR.

• 식물병연구
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• 제12권3호
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• pp.298-302
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• 2006

전형적인 줄무늬 모자이크 증상을 나타낸 칸나로부터 Cucumber mosaic virus(CMV)의 한 계통(Can-CMV)을 분리하고, 특성을 조사하였다. Can-CMV는 대부분의 전신감염 기주에서 병징이 발현되지 않았고, 이들 기주의 접종엽 및 상엽에서 RT-PCR로 바이러스의 감염여부를 조사한 결과, N. benthamiana와 N. glutinosa에서만 바이러스가 검출되었으며, 다른 기주로부터는 확인되지 않았다. 한편 Chenopodium amaranticolor의 접종엽에 발현된 국부 괴사병반은 대조로 접종한 Fny-CMV나 LS-CMV에 의해서 형성된 병반보다 매우 작은 크기의 반점을 형성하는 특성을 보였다. 또한 Vigna unguiculata의 접종엽에는 Fny-CMV나 LS-CMV가 접종 2-3일 후에 뚜렷한 괴사병반을 형성하는데 반해서, Can-CMV는 접종 4-5일 후에 윤곽이 확실치 않은 퇴록병반을 형성하였다. Can-CMV에 감염된 N. benthamiana로부터 추출한 dsRNA는 4종의 밴드가 검출되었으며, 이들 분자의 크기 및 종수는 Fny-CMV나 LS-CMV와 차이를 나타내지 않았다. Can-CMV의 항원은 Fny-CMV의 항혈청에 대해서 한 종의 뚜렷한 침강선을 형성하였으며, Fny-CMV의 항원에 의해서 형성된 침강선과 융합하였고, LS-CMV의 침강선과는 분지선을 형성함으로서, 혈청학적으로 서브그룹 I에 속하는 계통으로 판단되었다. 또한 Can-CMV에 감염된 N. benthamiana로부터 RNA를 추출하여 CMV-specific 프라이머를 이용한 CMV-RNA3의 외피단백질 유전자를 포함하는 3'영역에 대해서 RT-PCR을 실시하였다. Can-CMV는 Fny-CMV나 LS-CMV와 마찬가지로 예상되었던 약 950bp크기의 cDNA가 증폭되었으며, 이 cDNA를 EcoRI으로 처리하였을 때에는 절단되지 않았고, MspI에서는 595bp 및 350bp의 절편으로 절단되었다. 이와 같은 결과는 Fny-CMV의 패턴과 일치하였으며, 이러한 결과로부터 Can-CMV가 서브그룹 IA에 속하는 계통으로 확인되었다. 이상과 같은 결과들로부터, Can-CMV는 앞으로 바이러스와 기주의 다양한 상호관계를 이해하는데 있어서 중요한 병원학적 성질을 가지고 있는 것으로 생각되었다.

• 생명과학회지
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• 제17권2호통권82호
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• pp.198-203
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• 2007

형질전환은 유전자의 기능을 이해하는데 매우 중요한 기법이다. $Ca^{2+}$-인산 침전법은 시간과 비용이 저렴하여 가장 흔히 사용된다. 그러나 성숙 신경세포는 어린 신경세포나 다른 세포종에 비하여 형질전환이 어렵고 쉽게 죽는다. 본 연구에서는 Clontech사의 $CalPhos^{TM}$ Mammalian Transfection 방법을 수정하여 성숙한 신경세포를 효율적으로 형질전환할 수 있는 방법을 고안하였다. 대뇌 신경교세포를 DMEM/10% 말혈청에서 70-80% confluence까지 키우고 배지를 혈청이 첨가되지 않은 Neurobasal/Ara-C로 바꾸어 주어 더 이상 신경교세포가 분열하지 않게 한 다음, 여기에 E19 해마신경세포를 접종하여 배양하였다. $DNA/Ca^{2+}$-인산 침전물은 Clontech사의 $CalPhos^{TM}$ Mammalian Transfection Kit을 이용하여 크기($0.5-1\;{\mu}m$ in diameter) 및 농도(약 10 particles/$100\;{\mu}m^2$)를 배지에서 배양시간을 변화시켜 적당히 조절하였다. 이렇게 하면 in vitro에서 2주 이상 배양한 신경세포도 24-well plate 한 well당 10-15개의 형질전환된 건강한 신경세포를 얻을 수 있었다. 이 방법의 효용성을 검증하기 위하여 연접단백질인 $EGFP-CaMKII{\alpha}$ 융합단백질과 RFP 단백질 유전자(각각 $pEGFP-CaMKII{\alpha}$ 및 pDsRed2)를 형질전환한 결과 전자는 점박이 모양, 후자는 세포전체에 퍼진 양상의 표현을 관찰할 수 있었다. 따라서 본 연구는 성숙한 신경세포를 효율적으로 형질전환할 수 있는 방법을 제공한다.

• Asian-Australasian Journal of Animal Sciences
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• 제29권1호
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• pp.126-133
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• 2016

A gene from Actinomyces sp. Korean native goat (KNG) 40 that encodes an endo-${\beta}$-1,4-glucanase, EG1, was cloned and expressed in Escherichia coli (E. coli) $DH5{\alpha}$. Recombinant plasmid DNA from a positive clone with a 3.2 kb insert hydrolyzing carboxyl methyl-cellulose (CMC) was designated as pDS3. The entire nucleotide sequence was determined, and an open-reading frame (ORF) was deduced. The ORF encodes a polypeptide of 684 amino acids. The recombinant EG1 produced in E. coli $DH5{\alpha}$ harboring pDS3 was purified in one step using affinity chromatography on crystalline cellulose and characterized. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis/zymogram analysis of the purified enzyme revealed two protein bands of 57.1 and 54.1 kDa. The amino terminal sequences of these two bands matched those of the deduced ones, starting from residue 166 and 208, respectively. Putative signal sequences, a Shine.Dalgarno-type ribosomal binding site, and promoter sequences related to the consensus sequences were deduced. EG1 has a typical tripartite structure of cellulase, a catalytic domain, a serine-rich linker region, and a cellulose-binding domain. The optimal temperature for the activity of the purified enzyme was $55^{\circ}C$, but it retained over 90% of maximum activity in a broad temperature range ($40^{\circ}C$ to $60^{\circ}C$). The optimal pH for the enzyme activity was 6.0. Kinetic parameters, $K_m$ and $V_{max}$ of rEG1 were 0.39% CMC and 143 U/mg, respectively.

• BMB Reports
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• 제37권3호
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• pp.383-385
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• 2004

RNA interference (RNAi) is a process of sequence-specific gene silencing, which is initiated by double-stranded RNA (dsRNA). RNAi may also serve as an antiviral system in vertebrates. This study describes the inhibition of herpesvirus-6B (HHV-6B) replication by short interference RNAs (siRNAs) that are targeted to the U38 sequence that encodes DNA polymerase. When virus-infected SupT1 cells were treated by siRNA, these cells blocked the cytopathic effect (CPE) and detected the HHV-6B antibody-negative in indirect immunofluorescence assays (IFA). Our result suggests that RNAi can efficiently block Herpesvirus-6B replication.

• 대한한방내과학회지
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• 제23권2호
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• pp.306-312
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• 2002

Systemic Lupus Erythematosus(SLE) is a autoimmune disease characterized by combined symtoms of malar rash, discoid rash, neuropsychiatric disorder, renal disorder, hematologic disorder, photosensitivity immunologic disorder, oral ulcer, anti-nuclear antibody, arthritis, pleuritis and pericarditis, etc. Multiple genetic or environmental causes are supposed to facilitate antiboby production to autoantigen such as ds-DNA, histone, phospholipid, red blood cell, platelet, etc. And defective complementary system fail to remove autoantigen-antibody complex, which deposit in multiple organs and result in inflammatory damages. SLE does not correctly correspond to any specific category of oriental medicine. But, accoring to previous reports, it can be controlled by herb medications used differently patients-to-patients. So we are to report this one SLE case being successfully controlled by classic corticosteroids with herb medications based on oriental diffrential diagnosis of symptoms and signs.

• The Plant Pathology Journal
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• 제23권2호
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• pp.76-89
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• 2007

CaCDPK4, a full-length cDNA clone encoding Capsicum annuum calcium-dependent protein kinase 4, was isolated from chili pepper (Capsicum annuum L.). Deduced amino acid sequence of CaCDPK4 shares the highest homology with tobacco NpCDPK8 and chickpea CaCDPK2 with 79% identity. Genomic blot analyses revealed that CaCDPK4 is present as a single copy in pepper genome, but it belongs to a multigene family. CaCDPK4 was highly induced when pepper plants were inoculated with an incompatible bacterial pathogen. Induced levels of CaCDPK4 transcripts were also detected in pepper leaves by the treatment of ethephon, an ethylene-inducing agent, and high-salt stress condition. The bacterial-expressed GST-CaCDPK4 protein showed to retain the autophosphorylation activity in vitro. GUS expression driven by CaCDPK4 promoter was examined in transgenic Arabidopsis containing transcriptional fusion of CaCDPK4 promoter. GUS expression under CaCDPK4 promoter was strong in the root and veins of the seedlings. GW (-1965) and D3 (-1377) promoters conferred on GUS expression in response to inoculation of an incompatible bacterial pathogen, but D4-GUS (-913) and DS-GUS (-833) did not. Taken together, our results suggest that CaCDPK4 can be implicated on signal transduction pathway of defense response against an incompatible bacterial pathogen in pepper.

• 대한한의학회지
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• 제29권1호
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• pp.67-84
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• 2008

Objective : The main purpose of this study was to evaluate the effect of Jasinwhalhyul-tang (Zishenhuoxue-tang, JWT) on MRL/MpJ-Ipr/Ipr mouse model with systemic lupus erythematosus. Methods: The effect of JWT on MRL/MpJ-Ipr/Ipr mice that have autoimmune disease similar to SLE in humans was evaluated after JWT per oral in the present study. Mice were administered with Jasinwhalhyul-tang (Zishenhuoxue-tang, JWT) (80 or 400mg/kg) or distilled water for control group from experimental week 10 for 22 weeks. Results : The amount of erythematosus skin lesion and proteinuria were significantly decreased. The size and weight of cervical lymph nodes and spleen were significantly reduced. The ratio between activated $CD3^+CD69^+$ T-cells and undifferentiated $CD3^+CD4^-CD8^-$ T-cells in lymph nodes, spleen and kidney was effectively reduced. The gene expression of TGF-$\beta$ in spleen and kidney was increased. The amount of anti-dsDNA IgG in blood was decreased. The gene expression of TGF-$\beta$ in normal mouse spleen cells was increased depending on concentration by treatment of with T cell stimulating agent. In the histological examination of skin and kidney, the amount of infiltration of immune cells involved in the inflammatory response was decreased. Conclusions : According to the above results, JWT should be considered as an applicable therapeutic agent to SLE in clinical practice. Further research is required to investigate other efficacies of JWT on SLE.

• 한국약용작물학회지
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• 제22권5호
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• pp.398-404
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• 2014

Salt stress is one of the major abiotic stresses affecting the yield of ginseng (Panax ginseng C. A. Meyer). The objective of this study was to identify bio-marker, which is early responsive in salt stress in ginseng, using proteomics approach. Ginseng plants were exposed to 5 ds/m salt concentration and samples were harvested at 0, 6, 12 and 18 hours after exposure. Total proteins were extracted from ginseng leaves treated with salt stress using Mg/NP-40 buffer and were separated on high resolution 2-DE. Approximately $1003{\pm}240$ (0 h), $992{\pm}166$ (6 h), $1051{\pm}51$ (12 h) and $990{\pm}160$ (18 h) spots were detected in colloidal CBB stained 2D maps. Among these, 8 spots were differentially expressed and were identified by using MALDI-TOF/TOF MS or/and LC-MS/MS. Ethylene response sensor-1 (spot GL 1), nucleotide binding protein (spot GL 2), carbonic anhydrase-1 (spot GL 3), thylakoid lumenal 17.9 kDa protein (spot GL 4) and Chlorophyll a/b binding protein (spot GL 5, GL 6) were up-regulated at the 12 and 18 hour, while RuBisCO activase B (spot GL 7) and DNA helicase (spot GL 8) were down-regulated. Thus, we suggest that these proteins might participate in the early response to salt stress in ginseng leaves.

• 미생물학회지
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• 제54권1호
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• pp.77-78
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• 2018

포도상구균(Staphylococcus aureus)은 그람양성이고 구형의 박테리아로 Firmicutes 문에 속하며, 피부나 호흡기 감염 그리고 식중독의 주요 감염원 중에 하나이다. 포도상구균을 감염시키는 박테리오파지는 포도상구균 감염에 효과적인 처방으로 쓰일 수 있다. 본 연구에서는 충청남도에 위치한 가축 농장의 오수에서 분리된 포도상구균 박테리오파지 SA7 균주의 유전체 초안을 분석하였다. 본 균주는 G + C 비율이 34.1%이며, 34,730 bp 로 구성된 dsDNA 를 지니고 있었다. 염색체에서 53 개의 단백질 코딩 유전자가 확인되었으며, 이 중 23 개의 유전자는 BLASTp 분석으로부터 기능을 가지고 있다고 추정되었다. 나머지는 가설 단백질 혹은 보존 단백질이었다.