• 한국식물병리학회지
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• 제11권1호
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• pp.66-72
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• 1995

The coat protein (CP) gene of cucumber mosaic virus-As (CMV-As) strain was engineered for expression in the plant by using the cauliflower mosaic virus 35S transcript regulatory sequences. The CP gene was cloned into an Agrobacterium-derived binary vector. A chimeric gene was constructed by the cDNA of CMV-As CP and plant expression vector pBI121. The clone, pCMAS66, was first introduced into the phagemid vector pSPORT1 for situating sense orientation for translation and making restriction sites in order to re-introduce plant expression vector, pHI121. The resulting subclone pCASCP02 and plant expression vector pBI121 were treated with BamHI-SacI for excising the target gene and removing GUS gene, respectively. After Agrobacterium transformation by freeze-thaw technique, the clone, pCMASCP121-123 which contains sense orientation of the target gene, was selected and confirmed by restriction endonuclease analysis. The CMV-As CP gene was introduced into A. tumefaciens. The results on tobacco plant transformation with the vector system revealed that the system could be successfully introduced and showed high frequency of selection to putative transformations.

• Applied Biological Chemistry
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• 제38권1호
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• pp.49-54
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• 1995

한국 마늘에 감염된 바이러스의 종류와 병 발생 메카니즘을 구명하기 위하여, 마늘 바이러스 cDNA clone들을 분리하였다. 24개 cDNA clone들의 부분적인 염기 서열을 결정하였고, 이 중 poly(A) tail을 가진 5개 clone들의 염기 서열을 결정하였다. 이를 이미 알려진 다른 식물 바이러스와 비교했을 때, clone V9은 일차구조가 carlavirus와 유사성을 보이므로 GLV cDNA clone으로 여겨진다. Northern blot 결과로부터 GLV genome의 크기는 8.5 knt이고, poly(A) tail을 가지고 있다는 것을 알 수 있었다. clone V9의 3' 말단부분에는 바이러스 복제과정에서 cis-acting element로 작용한다고 여겨지는 hexanucleotide motif(5'-ACCUAA)가 존재한다. 또한 carlavirus의 껍질 단백질 subgenomic RNA의 5' 말단에 보존되어 있는 5'-TTAGGT도 나타난다. 이들은 모두 carlavirus의 특징들이다. 껍질 단백질 유전자를 pRSET-A 발현 벡터에 재조합하고, E. coli BL21에서 발현시켰다. 발현된 껍질 단백질을 $Ni^{2+}$ NTA affinity chromatography에 의해 정제하였다. 껍질 단백질을 토끼에 주사하여 항체를 만든 후, immunoblot을 한 결과 GLV 껍질 단백질에 해당하는 24 kDa polypeptide가 인지되었다. 또한 다양한 마늘 품종에 대해서 immunoblot을 한 결과, GLV 껍질 단백질의 크기와 GLV의 감염정도가 마늘 품종에 따라서 차이가 있다는 것을 알 수 있었다.

• Asian-Australasian Journal of Animal Sciences
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• 제27권11호
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• pp.1532-1539
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• 2014

Although growth rate is one of the main economic traits of concern in pig production, there is limited knowledge on its epigenetic regulation, such as DNA methylation. In this study, we conducted methyl-CpG binding domain protein-enriched genome sequencing (MBD-seq) to compare genome-wide DNA methylation profile of small intestine and liver tissue between fast- and slow-growing weaning piglets. The genome-wide methylation pattern between the two different growing groups showed similar proportion of CpG (regions of DNA where a cytosine nucleotide occurs next to a guanine nucleotide in the linear sequence) coverage, genomic regions, and gene regions. Differentially methylated regions and genes were also identified for downstream analysis. In canonical pathway analysis using differentially methylated genes, pathways (triacylglycerol pathway, some cell cycle related pathways, and insulin receptor signaling pathway) expected to be related to growth rate were enriched in the two organ tissues. Differentially methylated genes were also organized in gene networks related to the cellular development, growth, and carbohydrate metabolism. Even though further study is required, the result of this study may contribute to the understanding of epigenetic regulation in pig growth.

• 대한수의학회지
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• 제34권4호
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• pp.759-768
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• 1994

E coli strains isolated from pigs were investigated with respect to antimicrobial resistances and prevalence of R-plasmids. Also determined were properties of R-plasmids by plasmid conjugation, curing and southern hybridization using gene probes. All of 400 E coli strains were resistant to CL and SU, and 0.3% to 96.8% of the strains were resistant to most antimicrobials such as TC, PG, AM, SM, CP, GM, EM, NM, etc, while all strains were sensitive to AK. All strains were also multiply resistant to three to twelve antimicrobials. The resistances to PG, SM, TC, AM, CP, SU and ST were transferable and supposed to be mediated by R-plasmids which were opportunistic for transposition into chromosome. Plasmids bigger in size than chromosomal DNA were considered as R-plasmids and most plasmids in small size (<4Kb) proved as cryptic plasmids or nonconjugative R-plasmids. In a strain(No 99), AM resistant property was determined from both chromosomal DNA and R-plasmid DNA which is bigger in size than chromosome.

• Molecules and Cells
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• 제39권12호
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• pp.888-897
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• 2016

Stable expression of Foxp3 is ensured by demethylation of CpG motifs in the Foxp3 intronic element, the conserved non-coding sequence 2 (CNS2), which persists throughout the lifespan of regulatory T cells (Tregs). However, little is known about the mechanisms on how CNS2 demethylation is sustained. In this study, we found that Ten-Eleven-Translocation (Tet) DNA dioxygenase protects the CpG motifs of CNS2 from re-methylation by DNA methyltransferases (Dnmts) and prevents Tregs from losing Foxp3 expression under inflammatory conditions. Upon stimulation of Tregs by interleukin-6 (IL6), Dnmt1 was recruited to CNS2 and induced methylation, which was inhibited by Tet2 recruited by IL2. Tet2 prevented CNS2 re-methylation by not only the occupancy of the CNS2 locus but also by its enzymatic activity. These results show that the CNS2 methylation status is dynamically regulated by a balance between Tets and Dnmts which influences the expression of Foxp3 in Tregs.

• The Plant Pathology Journal
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• 제20권2호
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• pp.147-154
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• 2004

A simple and reliable procedure for RT-PCR detection of Apple stem pitting virus (ASPV), Cherry rasp leaf virus (CRLV), and Cherry necrotic rusty mottle virus (CNRMV) was developed. Two virus specific primer sets for each virus were found to specifically detect each virus among fourteen sets of designed oligonucleotide primers. Total RNAs extracted from healthy and from ASPV-,CRLV- and CNRMV-infected plant tissues were used to synthesize cDNA using oligo dT primer and then amplified by virus-specific primers for each virus. Each primer specifically amplified DNA fragments of 578 bp and 306 bp products for ASPV (prAS CP-C and prAS CP-N primers, respectively); 697 bp and 429 bp products for CRLV (prCR4 and prCR5-JQ3D3 primers, respectively); and 370 bp and 257 bp products for CNRMV (prCN4 and prCN6-NEG 1 primers, respec-tively) by RT-PCR. DNA sequencing of amplified DNA fragments confirmed the nature of each amplified DNA. Altogether, these results suggest that these virus specific primer sets can specifically amplify viral sequences in infected tissues and thus indicate that they can be used for specific detection of each virus.

• 식물분류학회지
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• 제48권4호
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• pp.289-300
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• 2018

Coreanomecon is a monotypic and endemic genus in Korea, distributed mainly in the southern regions. Coreanomecon is morphologically similar to Hylomecon by producing red latex, easily distinguished from Chelidonium, which produces yellow latex. Coreanomecon were merged into Hylomecon or Chelidonium depending on the authors. To understand the phylogenetic relationship of Coreanomecon, DNA sequences of chloroplast rbcL and matK and nuclear Internal Transcribed Spacer (ITS) regions were determined from the species of Papaveroideae (Papaveraceae) in Korea and analyzed with the Maximum Parsimony and Bayesian methods. Phylogenetic analyses of Papaveroideae suggest that Coreanomecon is sister to the clade of Chelidonium and Stylophorum in the ITS data and that it is sister to Hylomecon in the chloroplast (cpDNA) data. A constraining analysis using the Shimodaira-Hasegawa test (S-H test) suggested that the ITS data do not reject the sister relationship of Coreanomecon and Hylomecon. The S-H test also suggested that the cpDNA data is compatible with the placement of Coreanomecon as a sister to the clade of Chelidonium and Stylophorum. Although the conflicting phylogenetic results may stem from insufficient phylogenetic signals, they may also be associated with hybridization between Hylomecon and an ancestor of Stylophorum and Chelidonium. The results of this study suggest that Coreanomecon is a distinct lineage as an endemic genus, supporting the morphological data.

• Genomics & Informatics
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• 제15권1호
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• pp.28-37
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• 2017

Obesity is a highly prevalent, chronic disorder that has been increasing in incidence in young patients. Both epigenetic and genetic aberrations may play a role in the pathogenesis of obesity. Therefore, in-depth epigenomic and genomic analyses will advance our understanding of the detailed molecular mechanisms underlying obesity and aid in the selection of potential biomarkers for obesity in youth. Here, we performed microarray-based DNA methylation and gene expression profiling of peripheral white blood cells obtained from six young, obese individuals and six healthy controls. We observed that the hierarchical clustering of DNA methylation, but not gene expression, clearly segregates the obese individuals from the controls, suggesting that the metabolic disturbance that occurs as a result of obesity at a young age may affect the DNA methylation of peripheral blood cells without accompanying transcriptional changes. To examine the genome-wide differences in the DNA methylation profiles of young obese and control individuals, we identified differentially methylated CpG sites and investigated their genomic and epigenomic contexts. The aberrant DNA methylation patterns in obese individuals can be summarized as relative gains and losses of DNA methylation in gene promoters and gene bodies, respectively. We also observed that the CpG islands of obese individuals are more susceptible to DNA methylation compared to controls. Our pilot study suggests that the genome-wide aberrant DNA methylation patterns of obese individuals may advance not only our understanding of the epigenomic pathogenesis but also early screening of obesity in youth.

• Journal of Microbiology and Biotechnology
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• 제19권6호
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• pp.610-615
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• 2009

As a provirus, polydnavirus has a segmented DNA genome on chromosome(s) of host wasp. It contains several genes in each segment that presumably play critical roles in regulating physiological processes of target insect parasitized by the wasp. A cysteine-rich protein 1 (CRP1) is present in the polydnavirus Cotesia plutellae bracovirus (CpBV) genome, but its expression and physiological function in Plutella xylostella parasitized by the viral host C. plutellae is not known. This CpBV-CRP1 encoding 189 amino acids with a putative signal peptide (20 residues) was persistently expressed in parasitized P. xylostella with gradual decrease at the late parasitization period. Expression of CpBV-CRP1 was tissue-specific in the fat body/epidermis and hemocyte, but not in the gut. Its physiological function was analyzed by inducing transient expression of a CpBV segment containing CpBV-CRP1 and its promoter, which caused significant reduction in hemocyte -spreading and delayed larval development. When the treated larvae were co-injected with double-stranded RNA of CpBV-CRP1, the expression of CpBV-CRP1 disappeared, whereas other genes encoded in the CpBV segment was expressed. These co-injected larvae significantly recovered the hemocyte-spreading capacity and larval development rate. This study reports that CpBV-CRP1 is expressed in P. xylostella parasitized by C. plutellae and its physiological function is to alter the host immune and developmental processes.

• Parasites, Hosts and Diseases
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• 제55권2호
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• pp.115-120
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• 2017

Encystation mediating cyst specific cysteine proteinase (CSCP) of Acanthamoeba castellanii is expressed remarkably during encystation. However, the molecular mechanism involved in the regulation of CSCP gene expression remains unclear. In this study, we focused on epigenetic regulation of gene expression during encystation of Acanthamoeba. To evaluate methylation as a potential mechanism involved in the regulation of CSCP expression, we first investigated the correlation between promoter methylation status of CSCP gene and its expression. A 2,878 bp of promoter sequence of CSCP gene was amplified by PCR. Three CpG islands (island 1-3) were detected in this sequence using bioinformatics tools. Methylation of CpG island in trophozoites and cysts was measured by bisulfite sequence PCR. CSCP promoter methylation of CpG island 1 (1,633 bp) was found in 8.2% of trophozoites and 7.3% of cysts. Methylation of CpG island 2 (625 bp) was observed in 4.2% of trophozoites and 5.8% of cysts. Methylation of CpG island 3 (367 bp) in trophozoites and cysts was both 3.6%. These results suggest that DNA methylation system is present in CSCP gene expression of Acanthamoeba. In addition, the expression of encystation mediating CSCP is correlated with promoter CpG island 1 hypomethylation.