• Title/Summary/Keyword: reverse-transcription-PCR

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The Effect of Over-expression and Inactivation of Nuclear Factor I-C on the Dentin Matrix Gene Expression of MDPC-23 Odontoblasts (Nuclear Factor I-C 과발현과 발현억제가 MDPC-23 상아모세포주의 상아질 기질유전자 발현에 미치는 영향)

  • Bae, Hyun-Sook;Cho, Young-Sik
    • Journal of dental hygiene science
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    • v.9 no.4
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    • pp.427-433
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    • 2009
  • Nuclear factor I-C (NFI-C) null mice demonstrated aberrant odontoblast differentiation and abnormal dentin formation. In order to elucidate the mechanisms responsible for these changes, we evaluated the expression of dentin matrix gene after over-expression and inactivation of NFI-C in MDPC-23 cells by reverse transcription-polymerase chain reaction (RT-PCR) analysis. Collagen type I (Col I), osteocalcin (OC), and dentin sialophosphoprotein (DSPP) expression was decreased after inactivation of NFI-C. However, bone sialoprotein (BSP) expression was dramatically increased after inactivation of NFI-C. ALP and DMP4 expression was not changed after inactivation of NFI-C. The expression of alkaline phoshatase (ALP) and dentin matrix protein 4 (DMP4) was increased after over-expression of NFI-C, while Col I, OC, DSPP, and BSP expression was decreased. These findings suggest that odontoblasts after loss of NFI-C lost the phenotype of odontoblasts and acquired those of osteoblasts.

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Expression of MAGE A 1-6 and SSX 1-9 Genes in the Sputum and Cancer Tissue of the Lung Cancer Patients (폐암환자의 객담 및 암 조직에서 MAGE A 1-6와 SSX 1-9 유전자의 발현)

  • Lee, Yeun-Jae;Lee, Jang-Hoon;Lee, Jung-Cheul;Lee, Kwan-Ho
    • Tuberculosis and Respiratory Diseases
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    • v.70 no.4
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    • pp.315-322
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    • 2011
  • Background: A variety of diagnostic modalities for lung cancer have been developed. To achieve efficient and early detection of lung cancer, we tried to measure the expression rates of the melanoma associated gene (MAGE) and synovial sarcoma on X chromosome (SSX) genes. Methods: We designed primers for the SSX gene. In addition to the pre-developed MAGE A primer, using an SSX gene primer was attempted to increase the detection rate. We obtained cancer tissues and cancer-free lung tissues from resected lung, sputum from lung cancer patients who had not undergone surgery, and sputum from healthy people and patients with benign intrathoracic diseases. Results: The sensitivity of the MAGE or SSX gene RT-PCR to identifying cancer tissue of the 69 lung cancer patients was 95.2% for squamous cell carcinoma (scc), 87.0% for adenocarcinoma, and 100% for small cell carcinoma. The mean sensitivity value was 94.2% (p=0.001). For adenocarcinoma, the additional use of the SSX gene resulted in a higher expression rate than MAGE alone (87% vs. 69.6%). The expression rate for the cancer-free lung tissue was 14.3% in scc, 17.4% in adenocarcinoma, and 25.0% in small cell carcinoma. In the induced sputum of 49 lung cancer patients who had not undergone surgery, the expression rate for one of the two genes was 65.5%. The expression rate for the sputum of healthy people and benign intrathoracic diseases by MAGE or SSX gene reverse transcription polymerase chain reaction (RT-PCR) was 3.8% and 17.7%. Conclusion: Detecting lung cancer using the expression of MAGE and SSX genes in lung cancer tissue has high sensitivity.

Capsosiphon Fulvescens Decreases Melanin Synthesis Via Downregulation of Tyrosinase and TRP-2 Expression

  • Yoo, Hyun-Ju;Jo, Mi-Gyeong;Park, Si-Jun;Mun, Yeun-Ja;Pyo, Hyeong-Bae;Lim, Kyu-Sang;Lee, Ki-Nam;Woo, Won-Hong
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.21 no.4
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    • pp.998-1003
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    • 2007
  • The green marine algae, Capsosiphon fulvescens (CF) is a food supplement cultivated in south coast of Southern Korea. The purpose of this study was to investigate the mechanism of CF-induced hypopigmentation. The present study was designed to determine the effect of CF extracton melanogenesis in B16 cells, particularly its specific effects on tyrosinase and tyrosinase-related protein 2 (TRP-2). We measured melanin contents and analyzed melanosome associated protein levels using Western blot and Reverse transcription-polymerase chian reaction (RT-PCR) analysis. CF extract markedly inhibited melanin synthesis and tyrosinase activity. In addition, cellular dendricity was slightly decreased by CF extract. In further experiments, CF extract significantly reduced the protein levels of tyrosinase and TRP-2 in B16 cells. RT-PCR analysis revealed that tyrosinase and TRP-2 mRNA levels were unaffected by CF treatment. Therefore, these results suggest that hypopigmentary effect of CF was due to post-translational degradationof tyrosinase and TRP-2.

Cloning of Superoxide Dismutase (SOD) Gene of Lily 'Marcopolo' and Expression in Transgenic Potatoes

  • Park, Ji-Young;Kim, Hyun-Soon;Youm, Jung-Won;Kim, Mi-Sun;Kim, Ki-Sun;Joung, Hyouk;Jeon, Jae-Heung
    • Journal of Applied Biological Chemistry
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    • v.49 no.1
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    • pp.1-7
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    • 2006
  • Differential display reverse transcription PCR (DDRT-PCR) analysis was performed on lily 'Marcopolo' bulb scale for isolation of expressed genes during bulblet formation. Cu/Zn lily-superoxide dismutase (LSOD) of 872 bp gene, with ability to scavenge reactive oxygen in stress environment, was isolated. Northern blot analysis showed expression levels of LSOD maximized 12 days after bulblet formation. Ti plasmid vectors were constructed with sense and antisense expressions of LSOD gene and transformed into potato. Southern blot analysis of transgenic potatoes revealed different copies of T-DNA were incorporated into potato genome. In transgenic potatoes, lily SOD gene was overexpressed in sense lines and not in antisense lines. In native polyacrylamide gel electrophoresis analysis, additional engineered LSOD was detected in sense overexpressed transgenic line only. Transgenic potatoes were subjected to oxidative stress, such as herbicide methyl viologen (MV). Transgenic potato lines with sense orientation exhibited increased tolerance to MV, whereas in antisense lines exhibited decreased tolerance. In vitro tuberization of transgenic potato with sense orientation was promoted, but was inhibited in transgenic potato with antisense orientation.

Biological Evaluation of Bone Marrow-Derived Stem Cells onto Different Wettability by RT-PCR (역전사 중합효소 연쇄반응을 이용한 표면 적심성에 따른 골수유래 줄기세포의 생물학적 평가)

  • 김은정;박종수;김문석;조선행;이종문;이해방;강길선
    • Polymer(Korea)
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    • v.28 no.3
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    • pp.218-224
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    • 2004
  • The adhesion and proliferation of mammalian cells on polymeric biomaterials depend on the surface characteristics such as wettability, chemistry, charge and roughness. In order to recognize the correlation between the adhesion and proliferation of human bone marrow derived stem cells (BMSCs) and surface property, radio frequency generated plasma treatment on low density polyethylene (LDPE) has been carried out. The modified LDPE surfaces were characterized by measuring the static water contact angle. The adhesion and proliferation of cells on LDPE films were characterized by cell counting and reverse transcription-polymerase chain reaction (RT-PCR). The water contact angle of the film surface decreased with plasma treatment time. Proto-oncogenes (c-myc, c-fos) and tumor suppressor gene (p153) showed maximum expression with contact angle of 60 ∼ 70$^{\circ}$ range of LDPE film. By cell counting, we confirmed that the rate of cell proliferation appeared the higher on the film surface of the contact angle of 60∼70$^{\circ}$ We concluded that the surface wettability is an important role for the growth and differentiation of BMSCs.

First Report of Tobacco mild green mosaic virus Infecting Pepper in Korea

  • Choi, Gug-Seoun;Kim, Jae-Hyun;Ryu, Ki-Hyun;Choi, Jang-Kyung;Chae, Soo-Young;Kim, Jeong-Soo;Chung, Bong-Nam;Kim, Hyun-Ran;Choi, Yong-Mun
    • The Plant Pathology Journal
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    • v.18 no.6
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    • pp.323-327
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    • 2002
  • A rod-shaped virus was isolated from pepper showing mild mosic during the winter growing seasons of 2001 and 2002 in Korea. Based on its biological reactions, serological relationships, reverse transcription-poly-merase chain reaction (RT-PCR) using specific primers, and nucleotide sequence analysis of coat protein (CP) gene, the isolated virus was identified as Tobacco mild green mosaic virus (TMGMV) and designated as Korean pepper isolate (TMGMV-KP). Crude sap from infected tissue was mechanically transmitted to various indicator plants, which produced characteristic symptoms of tobamovirus infection. However, no symptom was observed in Gomphorena globosa. In RT-PCR assays with specific primers toy respective detection of TMGMV, Tobacco mosaic virus (TMV), Pepper mild mottle virue (PMMoV), and Tomato mosaic virus (ToMV), a single strong band of about 500 bp in length was produced from the sample used only with TMGMV primers. The amplified DNA was cloned and the nucleotide sequence was determined. Sequence comparisons with the CP gene of other tobamoviruses indicated that TMGMV-KP shared 99.3% identity with TMGMV Japanese isolate and only 59.1, 58.6, and 58.1% identity with TMV, PMMoV and ToMV, respectively. This is the first report of TMGMV in Korea.

The Detection of Plant Viruses in Korean Ginseng (Panax ginseng) through RNA Sequencing

  • Lee, Hong-Kyu;Kim, So-Yeon;Yang, Hee-Ji;Lee, Da-Som;Kwon, Boram;Lee, Dong-Yun;Oh, Jonghee;Lee, Su-Heon
    • The Plant Pathology Journal
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    • v.36 no.6
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    • pp.643-650
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    • 2020
  • Korean ginseng (Panax ginseng) is a dicotyledonous, medicinal, perennial plant belonging to the genus Panax of the family Araliaceae. We investigated the occurrence and incidence of plant viruses in Panax ginseng in Korea. A total of 656 leaf samples were combined into one and total RNA was extracted from the polled sample, using RNA sequencing (RNA-Seq), a metatranscriptome analysis of the plant virome was conducted. The virus present in Panax ginseng was confirmed by reverse transcription polymerase chain reaction (RT-PCR) assay using virus-specific primers. In RNA-Seq data analysis, the multiplication protein of four viral contigs including Aristotelia chilensis virus 1 (AcV1), Turnip mosaic virus (TuMV), Watermelon mosaic virus (WMV), and Tobamovirus multiplication protein were discovered. From our metatranscriptome analysis and RT-PCR assay, TuMV and WMV were detected, whereas the three viruses reported in China such as tomato yellow leaf curl China virus; panax notoginseng virus A; and panax virus Y were not found in this study. The distribution of domestic ginseng viruses seems different from that recorded in China. Overall, this is the first plant virome analysis of Panax ginseng in Korea.

Effects of Traditional Wine by using Mycelium of Phellinus Iinteus on the Expression of Inflammation-Related Proteins in HepG2 Cells (상황버섯 균사체를 이용한 전통주의 추출물이 HepG2 세포의 염증관련 단백질 발현에 미치는 영향)

  • Lee, Su-Jin;Han, Min-Ho;Lee, Yong-Tae;Huh, Man-Kyu;Chung, Kyung-Tae;Jeong, Young-Kee;Choi, Yung-Hyun;Choi, Byung-Tae
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.20 no.4
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    • pp.914-918
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    • 2006
  • It was examined that the effect of fermented traditional wine made by using mycelium of Phellinus linteus on the expression of inflammation-related proteins in HepG2 cells. HepG2 cells were incubated with or without ertract of traditional wine (ETMP), then analyzed by microscopic observation, reverse transcription polymerase chain reaction (RT-PCR) and Western blot. The results of RT-PCR and Western blot analyses showed that the level of inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2 and tumor necrosis factor $(TNF)-{\alpha}$ was induced by LPS, Dut the treatment of ETMP inhibited the expression of these proteins and its mRNAs. Besides, the results of Western blot analyses showed that the expression of nuclear $factor-{\kappa}Bp65$ and $inhibitory-{\kappa}B{\alpha}$ were also slightly affected by ETMP treatment. These results suggest that ETM P alleviate the expression of inflammation-related protein expressions and thus may be used as a functional alcoholic beverage.

Changes of cytokine mRNA expression and IgG responses In rats Infected with Capillaria hepatica

  • Kim, Dong-Kwan;Joo, Kyoung-Hwan;Chung, Myung-Sook
    • Parasites, Hosts and Diseases
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    • v.45 no.2 s.142
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    • pp.95-102
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    • 2007
  • The mRNA expression of several cytokines was evaluated in splenocytes and mesenteric lymph node (MLN) cells of rats infected with Capillaria hepatica by reverse-transcription (RT)-PCR until week 12 after infection. IgG1 and IgG2a, which are associated with Th1 and Th2 response, respectively, were also assessed by ELISA. The results indicated that the majority of cytokines, including the Th1 (IL-2 and $IFN-{\gamma}$ and Th2 cytokines (IL-4, IL-5 and IL-10) were expressed at maximal levels during the early stage of infection (after week 1-2), and the ELISA data also evidenced a similar pattern of changes in IgG1 arid IgG2a. Th1 and Th2 cytokines responded in a similar fashion in this rat model. The expression of cytokines in splenocytes was significantly higher than that in MLN cells, thereby indicating that cytokine production is controlled more by spleen than by MLN. In addition, the observation that $IFN-{\gamma}$ expression increased unexpectedly at the time of maximal egg production (6 weeks after infection) indicated that $IFN-{\gamma}$ is a cytokine reacting against egg production. However, increased IL-5 expression occurring in tandem with worm activity indicated that the activity of C. hepatica might be controlled by IL-5 expression.

The Effects of the 3-OH Group of Kaempferol on Interfollicular Epidermal Stem Cell Fate

  • Chae, Je Byeong;Choi, Hye-Ryung;Shin, Jung-Won;Na, Jung-Im;Huh, Chang-Hun;Park, Kyoung-Chan
    • Annals of dermatology
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    • v.30 no.6
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    • pp.694-700
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    • 2018
  • Background: Kaempferol (3,4',5,7-tetrahydroxyflavone) is a flavonoid known to have a wide range of pharmacological activities. The 3-OH group in flavonoids has been reported to determine antioxidant activities. Objective: We tested whether kaempferol can affect the expression of integrins and the stem cell fate of interfollicular epidermal stem cells. Methods: Skin equivalent (SE) models were constructed, and the expression levels of stem cell markers and basement membrane-related antigens were tested. The immunohistochemical staining patterns of integrins, p63, and proliferating cell nuclear antigen (PCNA) were compared between kaempferol- and apigenin-treated SE models. Reverse transcription-polymerase chain reaction (RT-PCR) was used to evaluate the mRNA expression of integrins. Results: Kaempferol increased the thickness of the epidermis when added to prepare SEs. In addition, the basal cells of kaempferol-treated SEs appeared more columnar. In the immunohistological study, the expression of integrins ${\alpha}6$ and ${\beta}1$ and the numbers of p63- and PCNA-positive cells were markedly higher in the kaempferol-treated model. However, apigenin showed no effects on the formation of three-dimensional skin models. RT-PCR analysis also confirmed that kaempferol increased the expression of integrin ${\alpha}6$ and integrin ${\beta}1$. Conclusion: Our findings indicated that kaempferol can increase the proliferative potential of basal epidermal cells by modulating the basement membrane. In other words, kaempferol can affect the fate of interfollicular epidermal stem cells by increasing the expression of both integrins ${\alpha}6$ and ${\beta}1$. These effects, in particular, might be ascribed to the 3-OH group of kaempferol.