• Journal of the korean academy of Pediatric Dentistry
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• v.32 no.1
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• pp.75-88
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• 2005

Ameloblastoma is the most commonly occurring odontogenic tumor in oral cavity. Although most are benign epithelial neoplasm, they are generally considered to be locally aggressive and destructive, exhibiting a high rate of recurrence. The biological behavior of this neoplasm is a slowly growing, locally invasive tumor without metastasis, therefore malignant neoplasm, changed its histological appearance to carcinoma or showed distant metastasis, is only defined clinically. In this study, we identified the differentially expressed genes(DEGs) in stages under benign or malignant ameloblastoma compared with normal patient using ordered differential display(ODD) reverse transcription(RT)-PCR and $GeneFishing^{TM}$ technology. ODD RT-PCR is rather effective when the investigation of samples containing very small amounts of total RNA must be accomplished. ODD RT-PCR used the means of amplification with anchored T-primer and adaptor specific primer. bearing definite two bases at their 3' ends and so this method could display differential 3'-expressed sequence taqs(ESTs) patterns without using full-length cDNAs. Compared with standard differential display, ODD RT-PCR is more simple and have enough sensitivity to search for molecular markers by comparing gene expression profiles, However, this method required much effort and skill to perform. $GeneFishing^{TM}$ modified from DD-PCR is an improved method for detecting differentially expressed genes in two or more related samples. This two step RT-PCR method uses a constant reverse primer(anchor ACP-T) to prime the RT reaction and arbitrary primer pairs(annealing control primers, ACPs) during PCR. Because of high annealing specificity of ACPs than ODD RT-PCR, the application of $GeneFishing^{TM}$ to DEG discovery generates reproducible, authentic, and long(100bp to 2kb) PCR products that are detectable on agarose gels. Consequently, various DEGs observed differential expression levels on agarose gels were isolated from normal, benign, and malignant tissues using these methods. The expression patterns of the some isolated DEGs through ODD RT-PCR and $GeneFishing^{TM}$ were confirmed by Northern blot analysis and RT-PCR. The results showed that these identified DEGs were implicated in ameloblastoma neoplasm processes. Therefore, the identified DEGs will be further studied in order to be applied in candidate selection for marker as an early diagnosis during ameloblastoma neoplasm processes.

• Molecular & Cellular Toxicology
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• v.1 no.4
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• pp.230-236
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• 2005

Formaldehyde is a common environmental contaminant found in tobacco smoke, paint, garments, diesel and exhaust, and medical and industrial products. Formaldehyde has been considered to be potentially carcinogenic, making it a subject of major environmental concern. However, only a little information on the mechanism of immunological sensitization and asthma by this compound has been known. So, we performed with Jurkat cell line, a human T lymphocyte, to assess the induction of DNA damage and to identify the DEGs related to immune response or toxicity by formaldehyde. In this study, we investigated the induction of DNA single strand breaks by formaldehyde using single cell gel electrophoresis assay (comet assay). And we compared gene expression between control and formaldehyde treatment to identify genes that are specifically or predominantly expressed by employing annealing control primer (ACP)-based $GeneFishing^{TM}$ method. The cytotoxicity ($IC_{30}$) of formaldehyde was determined above the 0.65 mM in Jurkat cell in 48 h treatment. Based on the $IC_{30}$ value from cytotoxicity test, we performed the comet assay in this concentration. From these results, 0.65 mM of formaldehyde was not revealed significant DNA damages in the absence of S-9 metabolic activation system. And the one differentially expressed gene (DEG) of formaldehyde was identified to zinc finger protein 292 using $GeneFishing^{TM}$ method. Through further investigation, we will identify more meaningful and useful DEGs on formaldehyde, and then can get the information on the associated mechanism and pathway with immune response or other toxicity by formaldehyde exposure.

• Archives of Plastic Surgery
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• v.34 no.6
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• pp.671-678
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• 2007

Purpose: Venous malformation(VM) which often causes pain and discomfort is the most common type of vascular malformations. Although it is presented with disfigured appearance and associated soft tissue or skeletal hypertrophy, the molecular bases of VMs are poorly understood. Differentially expressed genes(DEGs) of VMs were investigated to illuminate the molecular mechanism of the disease entity. Methods: Gene expressions of VM patients' subcutaneous tissue were studied in comparison with normal persons' by $GeneFishing^{TM}$ technique using the annealing control primers (ACPs) to identify DEGs. Candidate genes were sequenced and screened by basic local alignment search tool (BLAST) afterwards. Results: Among seventy DEGs identified, forty DEGs which had shown significantly different expression pattern were sequenced. Twenty eight out of 40 were up-regulated while 12 were down-regulated. BLAST searches revealed that 37 were known genes and 3 were unknown genes. Many genes were involved in the differentiation and remodeling of smooth muscle cells, opposed to the previous hypothesis that a lot of angiogenetic genes would be involved. Furthermore, several transcription factors and related genes, as well as cell signaling and metabolism regulators, were up regulated. Conclusion: It suggests that analysis of DEGs in VMs provide basic knowledge about its pathophysiology. and new therapeutic approaches.

• Reproductive and Developmental Biology
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• v.31 no.1
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• pp.55-60
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• 2007

Aging causes thymus involution, and genes in thymus play an important role in the development of the immune system. In this study, we compared genes expressed in thymus of neonatal and peripubertal rats using annealing control primers (ACPs)-based GeneFishing polymerase chain reaction (PCR) and semiquantitative reverse transcription (RT)-PCR. We identified 10 differentially expressed genes (DEGs) with 20 ACPs. Of 10 DEGs, bystin-like, collagen type V alpha 1 (COL5A1), and T-cell receptor beta-chain segment 2 (TCRB2) that are related to immune-function were detected in rat thymus. Bystin-like and TCRB2 were up-regulated, while COL5A1 was down-regulated in peripubertal thymus. Semiquantitative RT-PCR confirmed postnatal changes in expression of bystin-like, COL5A1, and TCRB2. These results suggest that bystin-like, COL5A1, and TCRB2 could regulate immune function controlled in thymus as age increases.

• Molecular & Cellular Toxicology
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• v.4 no.1
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• pp.22-30
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• 2008

Malachite Green (MG), a toxic chemical used as a dye, topical antiseptic and antifungal agent for fish, is highly soluble in water, cytotoxic to various mammalian cells and also acts as a liver tumor promoter. In view of its industrial importance and possible exposure to human beings, MG possesses a potential environmental health hazard. So, we performed with HepG2, a human hepatocellular carcinoma cell line, to identify the differentially expressed genes (DEGs) related to toxicity of MG. And we compared gene expression between control and MG treatment to identify genes that are specifically or predominantly expressed by employing annealing control primer (ACP)-based $GeneFishing^{TM}$ method. The cytotoxicity $(IC_{20})$ of MG was determined above the $0.867{\mu}M$ in HepG2 cell for 48 h treatment. And the DEGs of MG were identified that 5 out of 6 DEGs were upregulated and 1 out of 6 DEGs was down-regulated by MG. Also, MG induced late apoptosis and necrosis in a dose dependent in flow cytometric analysis. Through further investigation, we will identify more meaningful and useful DEGs on MG, and then can get the information on mechanism and pathway associated with toxicity of MG.

• The Korean Journal of Physiology and Pharmacology
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• v.14 no.5
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• pp.265-272
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• 2010

Follicular cystic ovary (FCO) is one of the most frequently diagnosed ovarian diseases and is a major cause of reproductive failure in mammalian species. However, the mechanism by which FCO is induced remains unclear. Genetic alterations which affect the functioning of many kinds of cells and/or tissues could be present in cystic ovaries. In this study, we performed a comparison analysis of gene expression in order to identify new molecules useful in discrimination of bovine FCO with follicular cystic follicles (FCFs). Normal follicles and FCFs were classified based on their sizes (5 to 10 mm and $\geq25mm$). These follicles had granulosa cell layer and theca interna and the hormone $17{\beta}$-estradiol ($E_2$)/ progesterone ($P_4$) ratio in follicles was greater than one. Perifollicular regions including follicles were used for the preparation of RNA or protein. Differentially expressed genes (DEG) that showed greater than a 2-fold change in expression were screened by the annealing control primer (ACP)-based PCR method using $GeneFishing^{TM}$ DEG kits in bovine normal follicles and FCFs. We identified two DEGs in the FCFs: ribosomal protein L15 (RPL15) and microtubule-associated protein 1B (MAP1B) based on BLAST searches of the NCBI GenBank. Consistent with the ACP analysis, semi-quantitative PCR data and Western blot analyses revealed an up-regulation of RPL15 and a down-regulation of MAP1B in FCFs. These results suggest that RPL15 and MAP1B may be involved in the regulation of pathological processes in bovine FCOs and may help to establish a bovine gene data-base for the discrimination of FCOs from normal ovaries.

• Journal of Crop Science and Biotechnology
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• v.10 no.4
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• pp.231-236
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• 2007

Differentially expressed genes(DEG) were identified in a rice variety, Sathi, an indica type showing high allelopathic potential against barnyardgrass(Echinochloa crus-galli(L.) Beauv. var. frumentaceae). Rice plants were grown with and without barnyardgrass and total RNA was extracted from rice leaves at 45 days after seeding. DEG full-screening was performed by $GeneFishing^{TM}$ method. The differentially expressed bands were re-amplified and sequenced, then analyzed by Basic Local Alignment Search Tool(BLAST) searching for homology sequence identification. Gel electrophoresis showed nine possible genes associated with allelopathic potential in Sathi, six genes(namely DEG-1, 4, 5, 7, 8, and 9) showed higher expression, and three genes(DEG-2, 3 and 6) showed lower expression as compared to the control. cDNA sequence analysis showed that DEG-7 and DEG-9 had the same sequence. From RT PCR results, DEG-6 and DEG-7 were considered as true DEG, whereas DEG-1, 2, 3, 4, 5, and 8 were considered as putative DEG. Results from blast-n and blast-x search suggested that DEG-1 is homologous to a gene for S-adenosylmethionine synthetase, DEG-2 is homologous to a chloroplast gene for ribulose 1,5-bisphosphate carboxylase large subunit, DEG-8 is homologous to oxysterol-binding protein with an 85.7% sequence similarity, DEG-5 is homologous to histone 2B protein with a 47.9% sequence similarity, DEG-6 is homologous to nicotineamine aminotransferase with a 33.1% sequence similarity, DEG-3 has 98.8% similarity with nucleotides sequence that has 33.1% similarity with oxygen evolving complex protein in photosystem II, DEG-7 is homologous to nucleotides sequence that may relate with putative serin/threonine protein kinase and putative transposable element, and DEG-4 has 98.8% similarity with nucleotides sequence for an unknown protein.

• Journal of The Korean Society of Grassland and Forage Science
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• v.34 no.2
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• pp.114-119
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• 2014

Salinity and drought stresses are probably the most significant abiotic factor limiting plant's growth, also negatively affect seed germination and early seedling development. To study on effect of NaCl and PEG stress on seed germination and gene expression pattern of tall fescue, the levels of NaCl and PEG-induced water stresses were determined in first experiment. Different concentration of NaCl (0 to 350 mM) and PEG (0 to 30%) were used for seed treatment. Seed Germination percentage reduced with increasing osmotic potential of growth medium either due to NaCl or PEG. Seeds were not germinate at 350 mM NaCl or 30% PEG treatment. On the basis of the results, Kentucky31(E-) had more resistant than Fawn in both stress conditions. Furthermore, we have used an annealing control primer-based differential display reverse transcription-polymerase chain reaction method to identify salt- and drought stress-induced differentially expressed genes (DEGs) in tall fescue leaves. Using 120 annealing control primers, a total of 4 genes were identified and sequenced. The possible roles of the identified DEGs are discussed in the context of their putative role during salinity and drought stresses.

• Journal of Korean Neurosurgical Society
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• v.48 no.1
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• pp.20-30
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• 2010

Objective : We determined whether the expression of GRIM-19 is correlated with pathologic types and malignant grades in gliomas, and determined the function of GRIM-19 in human gliomas. Methods : Tumor tissues were isolated and frozen at $-80^{\circ}C$ just after surgery. The tissues consisted of normal brain tissue (4), astrocytomas (2), anaplastic astrocytomas (2), oligodendrogliomas (13), anaplastic oligodendrogliomas (11), and glioblastomas (16). To profile tumor-related genes, we applied RNA differential display using a $Genefishing^{TM}$ DEG kit, and validated the tumor-related genes by reverse transcription polymerase chain reaction (RT-PCR). A human glioblastoma cell line (U343MG-A) was used for the GRIM-19 functional studies. The morphologic and cytoskeletal changes were examined via light and confocal microscopy. The migratory and invasive abilities were investigated by the simple scratch technique and Matrigel assay. The antiproliferative activity was determined by thiazolyl blue Tetrazolium bromide (MTT) assay and FACS analysis. Results : Based on RT-PCR analysis, the expression of GRIM-19 was higher in astrocytic tumors than oligodendroglial tumors. The expression of GRIM-19 was higher in high-grade tumors than low-grade tumors or normal brain tissue; glioblastomas showed the highest expression. After transfection of GRIM-19 into U343MG-A, the morphology of the sense-transfection cells became larger and more spindly. The antisensetransfection cells became smaller and rounder compared with wild type U343MG-A. The MTT assay showed that the sense-transfection cells were more sensitive to the combination of interferon-$\beta$ and retinoic acid than U343MG-A cells or antisense-transfection cells; the antiproliferative activity was related to apoptosis. Conclusion : GRIM-19 may be one of the gene profiles which regulate cell death via apoptosis in human gliomas.