• Genomics & Informatics
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• 제18권3호
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• pp.26.1-26.6
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• 2020

Single-cell RNA sequencing (scRNA-seq) has been widely applied to provide insights into the cell-by-cell expression difference in a given bulk sample. Accordingly, numerous analysis methods have been developed. As it involves simultaneous analyses of many cell and genes, efficiency of the methods is crucial. The conventional cell type annotation method is laborious and subjective. Here we propose a semi-automatic method that calculates a normalized score for each cell type based on user-supplied cell type-specific marker gene list. The method was applied to a publicly available scRNA-seq data of mouse cardiac non-myocyte cell pool. Annotating the 35 t-stochastic neighbor embedding clusters into 12 cell types was straightforward, and its accuracy was evaluated by constructing co-expression network for each cell type. Gene Ontology analysis was congruent with the annotated cell type and the corollary regulatory network analysis showed upstream transcription factors that have well supported literature evidences. The source code is available as an R script upon request.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2005년도 창립 30주년 및 춘계학술대회
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• pp.18-18
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• 2005

• 한국가축번식학회지
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• 제27권3호
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• pp.225-231
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• 2003

본 연구는 생체 내 세포 및 조직배양기로서의 면역결핍동물을 개발할 목적으로 proximal lck promoter와 DT-A gene를 이용하여 형질전환생쥐을 생산하고 이 형질전환생쥐의 면역세포에서 DT-A gene이 발현되는지를 분석하였다. 형질전환생쥐와 정상생쥐로부터 thymus, spleen 및 liver에서 RNA를 추출하여 RT-PCR수행하였는데 정상생쥐의 조직에서는 어떠한 DT-A gene의 발현양상을 얻을 수 없었으나 형질전환생쥐의 thymus, spleen, liver에 DT gene의 발현을 확인할 수 있었고, Northern blotting을 이용하여 형질전환생쥐의 thymus, spleen 및 liver에서 DT-A gene이 강하게 발현되는 것으로 나타났다. 형질전환생쥐 $F_1$ 및 $F_2$ 산자의 혈액에서 T-cell 발달의 분포도를 확인하기 위해 CD4 및 CD8 antibody를 이용하여 FACS analysis를 실시하였는데 형질전환생쥐의 혈액 내 mature T-cell인 single positive thymocyte의 수가 정상생쥐에 비해 감소하는 경향을 나타냈다. 정상생쥐의 혈액 내 T-cell 중 $CD8^{+}$ T-cell의 경우 약 50%를 나타냈으나 형질전환생쥐의 경우 33%로 감소하였고, $CD4^{+}$ T-cell은 정상생쥐에서 10%를 차지하고 있으나 형질전환생쥐에서는 5.9%로 감소되는 것으로 분석되었다. 그러므로 본 연구의 형질전환생쥐에서 lck promoter에 의해 초기 immature한 상태의 T-cell에서 DT-A gene이 발현되어 발육중인 T-cell이 파괴 되어 mature 상태인 $CD4^{+}CD8^{-}$나 $CD4^{-}CD8^{+}$ cells (single-positive)들이 감소된 것으로 확인되었다.

• Plant Biotechnology Reports
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• 제4권1호
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• pp.75-83
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• 2010

Glycine betaine has been reported as an osmoprotectant compound conferring tolerance to salinity and osmotic stresses in plants. We previously found that the expression of betaine aldehyde dehydrogenase 1 gene (OsBADH1), encoding a key enzyme for glycine betaine biosynthesis pathway, showed close correlation with salt tolerance of rice. In this study, the expression of the OsBADH1 gene in transgenic tobacco was investigated in response to salt stress using a transgenic approach. Transgenic tobacco plants expressing the OsBADH1 gene were generated under the control of a promoter from the maize ubiquitin gene. Three homozygous lines of $T_2$ progenies with single transgene insert were chosen for gene expression analysis. RT-PCR and western blot analysis results indicated that the OsBADH1 gene was effectively expressed in transgenic tobacco leading to the accumulation of glycine betaine. Transgenic lines demonstrated normal seed germination and morphology, and normal growth rates of seedlings under salt stress conditions. These results suggest that the OsBADH1 gene could be an excellent candidate for producing plants with osmotic stress tolerance.

• Molecular & Cellular Toxicology
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• 제1권3호
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• pp.203-208
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• 2005

Thioacetamide (TA) is potent haptotoxincant that requires metabolic activation by mixed-function oxidases. Micrcarray technology, which is massive parallel gene expression profiling in a single hybridization experiment, has provided as a powerful molecular genetic tool for biological system related toxicant. In this study we focus on the use of toxicogenomics for the determination of gene expression analysis associated with hepatotoxicity in rat liver epithelial cell line WB-F344 (WB). The WB cells was used to assess the toxic effects of TA. WB cells were exposed to two concentrations of TA-doses which caused 20% and 50% cell death were chosen and the cells exposed for periods of 2 and 24 h. Our data revealed that following the 2-h exposure at the both of doses and 24-h exposure at the low doses, few changes in gene expression were detected. However, after 24-h exposure of the cells to the high concentration, multiple changes in gene expression were observed. TA treatment gave rise predominantly to up-regulation of genes involved in cell cycle and cell death, but down-regulation of genes involves in cell adhesion and calcium ion binding. Exposure of WB cells to higher doses of the TA gave rise to more changes in gene expression at lower exposure times. These results show that TA regulates expression of numerous genes via direct molecular signaling mechanisms in liver cells.

• BMB Reports
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• 제46권2호
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• pp.65-72
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• 2013

In situ detection of RNAs is becoming increasingly important for analysis of gene expression within and between intact cells in tissues. International genomics efforts are now cataloging patterns of RNA transcription that play roles in cell function, differentiation, and disease formation, and they are demon-strating the importance of coding and noncoding RNA transcripts in these processes. However, these techniques typically provide ensemble averages of transcription across many cells. In situ hybridization-based analysis methods complement these studies by providing information about how expression levels change between cells within normal and diseased tissues, and they provide information about the localization of transcripts within cells, which is important in understanding mechanisms of gene regulation. Multi-color, single-molecule fluorescence in situ hybridization (smFISH) is particularly useful since it enables analysis of several different transcripts simultaneously. Combining smFISH with immunofluorescent protein detection provides additional information about the association between transcription level, cellular localization, and protein expression in individual cells.

• Biotechnology and Bioprocess Engineering:BBE
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• 제6권2호
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• pp.117-127
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• 2001

The high-level expression of a human single chain urokinase-type plasminogen activator (scu-PA) was achieved by employing a methotrexate (MTX)-dependent gene amplification system in Chinese hamster ovary (CHO) cells. By cotransfecting and coamplifying a scu-PA expression plasmid and dihydrofolate reductase (DHFR) minigene, several scu-PA expressing CHO cell lines were selected and gene-amplified. These recombinant cell lines, NGpUKs, secreted a completely processed scu-PA of 54 kD and up to 60mg/L was accumulated in the culture medium when they were adapted to an optimal MTX concentration. Over 95% of the scu-PA expressed was secreted in the culture medium and identified having the proper function of a plasminogen activator when activated by plasmin. Based on a genomic Southern analysis, a representative subclone, MGpUK-5, exhibited MTX-dependent scu-PA gene amplification, plus the initial single-copy gene of scu-PA eventually turned into about 150 copies of the amplified gene of scu-PA after gradual adaptation to 2.0$\mu$M of MTX. Meanwhile, the transcripts kof the scu-PA gene increased, although -early saturation of transcription was identified at 0.1$\mu$M of MTX. The scu-PA production by the MGpUK-5 subclone also increased relative to the gene amplification and increased transcripts, however, the relationship was not linearly proportional. Accordingly, since the MGpUK cell lines expressed elevated levels of enzymatically active scu-PA, these cell lines could be applied to the largescale production of scu-PA.

• The Korean Journal of Physiology and Pharmacology
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• 제28권5호
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• pp.403-411
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• 2024

The global spread of flaviviruses has triggered major outbreaks worldwide, significantly impacting public health, society, and economies. This has intensified research efforts to understand how flaviviruses interact with their hosts and manipulate the immune system, underscoring the need for advanced research tools. RNA-sequencing (RNA-seq) technologies have revolutionized our understanding of flavivirus infections by offering transcriptome analysis to dissect the intricate dynamics of virus-host interactions. Bulk RNA-seq provides a macroscopic overview of gene expression changes in virus-infected cells, offering insights into infection mechanisms and host responses at the molecular level. Single-cell RNA sequencing (scRNA-seq) provides unprecedented resolution by analyzing individual infected cells, revealing remarkable cellular heterogeneity within the host response. A particularly innovative advancement, virus-inclusive single-cell RNA sequencing (viscRNA-seq), addresses the challenges posed by non-polyadenylated flavivirus genomes, unveiling intricate details of virus-host interactions. In this review, we discuss the contributions of bulk RNA-seq, scRNA-seq, and viscRNA-seq to the field, exploring their implications in cell line experiments and studies on patients infected with various flavivirus species. Comprehensive transcriptome analyses from RNA-seq technologies are pivotal in accelerating the development of effective diagnostics and therapeutics, paving the way for innovative treatments and enhancing our preparedness for future outbreaks.

• Molecules and Cells
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• 제42권3호
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• pp.189-199
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• 2019

Cell-to-cell variability in gene expression exists even in a homogeneous population of cells. Dissecting such cellular heterogeneity within a biological system is a prerequisite for understanding how a biological system is developed, homeostatically regulated, and responds to external perturbations. Single-cell RNA sequencing (scRNA-seq) allows the quantitative and unbiased characterization of cellular heterogeneity by providing genome-wide molecular profiles from tens of thousands of individual cells. A major question in analyzing scRNA-seq data is how to account for the observed cell-to-cell variability. In this review, we provide an overview of scRNA-seq protocols, computational approaches for dissecting cellular heterogeneity, and future directions of single-cell transcriptomic analysis.

• 한국생물정보학회:학술대회논문집
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• 한국생물정보시스템생물학회 2005년도 BIOINFO 2005
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• pp.161-164
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• 2005

Data mining techniques can be applied to identify patterns of interest in the gene expression data. One goal in mining gene expression data is to determine how the expression of any particular gene might affect the expression of other genes. To find relationships between different genes, association rules have been applied to gene expression data set [1]. A notable limitation of association rule mining method is that only the association in a single profile experiment can be detected. It cannot be used to find rules across different condition profiles or different time point profile experiments. However, with the appearance of time-series microarray data, it became possible to analyze the temporal relationship between genes. In this paper, we analyze the time-series microarray gene expression data to extract the sequential patterns which are similar to the association rules between genes among different time points in the yeast cell cycle. The sequential patterns found in our work can catch the associations between different genes which express or repress at diverse time points. We have applied sequential pattern mining method to time-series microarray gene expression data and discovered a number of sequential patterns from two groups of genes (test, control) and more sequential patterns have been discovered from test group (same CO term group) than from the control group (different GO term group). This result can be a support for the potential of sequential patterns which is capable of catching the biologically meaningful association between genes.