• The Journal of Internal Korean Medicine
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• v.24 no.1
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• pp.134-143
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• 2003

Objectives : We performed this study to understand the molecular basis of the antitumor effect of Saussurea lappa, Pharbitis nil, Plantago asiatica and Taraxacum mongolicum, which have been used for cancer treatment in Korean traditional medicine. Design: We analyzed, the effect of these medicinal herbs on proliferation and apoptosis of tumor cells and its association with gene expression, We performed semi-quantitative reverse transcription-polymerase chain reaction(RT-PCR) analysis of cell cycle- and apoptosis-related genes using a gastric cancer cell line AGS. Results : Cell counting assay and $[^3H]thymidine$ uptake analysis showed that Saussurea lappa and Pharbitis nil strongly inhibit cell proliferation of AGS in a dose-dependent manner. Interestingly, gene espression assay revealed that mRNA espression levels of c-Jun, c-Fos, c-Myc, and Cyclin D1 were markedly decreased by Saussurea lappa and Pharbitis nil. Furthermore, Saussurea lappa was identified to activate expression of the p53 tumor suppressor and its downstream effector $p21^{Wafl}$, which leads to $G_1$ cell cycle arrest and apoptosis. These observations suggest that the anticancer effect of Saussurea lappa and Pharbitis nil might be associated with their regulatory capability of tumor-related gene expression.

• Journal of Physiology & Pathology in Korean Medicine
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• v.16 no.3
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• pp.602-608
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• 2002

To investigate whether Puerariae radix (PR) possesses protective effect against ethanol (EtOH)-inducecl apoptosis in the central nervous system, MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, flow cytometric analysis, DNA fragmentation assay. and reverse transcription-polymerase chain reaction (RT-PCR) were performed on human neuroblastoma cell line SK-N-MC. Morphological and biochemical analyses demonstrated that SK-N-MC cells treated with EtOH exhibit classical apoptotic features. On the other hand, cells pre-treated with PR prior to EtOH exposure showed decreased occurrence of classical apoptotic features. In addition, it was shown that PR pre-treatment inhibits EtOH-induced increases in the levels of mRNA expression of bax and caspase-3, while it further enhances the level of bcl-2 expression. These results suggest that PR may exert protective effects against EtOH-induced apoptosis in human neuroblastoma cells.

• Journal of Physiology & Pathology in Korean Medicine
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• v.22 no.3
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• pp.678-683
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• 2008

Aconitum pseudo-laeve var. erectum has traditionally been used for the treatment of water retention in the body. Administration of the aqueous extract of Aconitum pseudo-laeve var. erectum has the efficiency of anti-inflammatory activity and modulates the intestinal immune system. However, the mechanism of anti-inflammatory action of Aconitum pseudo-laeve var. erectum has not been clarified yet. In the present study, the effect of Aconitum pseudo-laeve var. erectum against LPS-stimulated expressions of COX-2 and iNOS in cells of the murine RAW 264.7 macrophages was investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, reverse transcription- polymerase chain reaction (RT-PCR), PGE2 immunoassay, and NO detection. The results of the present study indicate that Aconitum pseudo-laeve var. erectum is a potent inhibitor of the LPS-induced NO and $PGE_{2}$ production by blocking iNOS and $NF{\kappa}B$ activation in RAW 264.7 macrophages. These findings suggest that Aconitum pseudo-laeve var. erectum is a potential therapeutic for the treatment of inflammatory syndrome.

• Journal of Plant Biotechnology
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• v.43 no.1
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• pp.37-48
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• 2016

The transcriptional profiles of 'Tamnara' grapevine (Vitis labruscana L.) to Rhizobium vitis were determined using 12,000 gene oligonucleotide microarray chips constructed with 6,776 unigenes based on the EST sequencing. Among them, 95 clones were up-regulated more than three times and 90 were down-regulated more than 5-times in the R. vitis-inoculated grapevines relative to the control vines. Treatment of salicylic acid showed that 337 clones were upregulated and 52 clones were down regulated in grapevines. Microarray analysis, reverse transcription-polymer chain reaction, and slot blot hybridization analysis revealed that 5, 14, and 64 clones were up-regulated and 10, 12, and 61 clones were down-regulated in wounded, salicylic acid-treated, and R. vitis-inoculated 'Tamnara' grapevine leaves, respectively. The expression patterns of ${\beta}$-1,3-glucanase, proline-rich protein, and lipoxygenase genes of 'Tamnara' moderately resistant to R. vitis were similar to those of resistant 'Concord' and 'Delaware' grapevines. However, chalcone synthase genes in 'Tamnara' grapevines showed similar expression patterns to susceptible grapevines 'Neomuscat' and 'Rizamat'. Further expression studies with various clones for each gene should be conducted to elucidate their roles in resistant responses against pathogens or other stimuli in grapevines. These results could provide better resources for understanding the mechanism of defense responses against crown gall disease and clues for identifying new genes that may play a role in defense against R. vitis in grapevines.

• Korean Journal of Veterinary Research
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• v.46 no.4
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• pp.355-361
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• 2006

Porcine epidemic diarrhea virus (PEDV) strain Chinju99, which was previously isolated from piglets suffering from severe diarrhea was used to characterize the membrane (M) protein gene to establish the molecular information, and the results will be useful in elucidating concepts related to molecular pathogenesis and antigenic structures of PEDV isolates. The Chinju99 M gene generated by reverse transcription and polymerase chain reaction (RT-PCR) consisted of 681 bases containing 22.3% adenine, 22.3% cytosine, 23.1% guanine and 32.3% thymine nucleotides, and the GC content was 45.4%. It had some nucleotide mismatches from M gene of other PEDV strains, such as CV777, Br1/87, KPEDV-9, JMe2, JS2004-2 and LJB-03 with 97-99% nucleotide sequence homology to these strains. Also, it encoded a protein of 226 amino acids, which had some mismatches from those of CV777, Br1/87, KPEDV-9, JMe2, JS20004-2 and LJB-03, as the amino acid sequence homology showed a 97-98% to these strains. The Chinju99 had a very close relationship to the Japanese strain JMe2 for the nucleotide and amino acid sequences of the M gene. The amino acids predicted from Chinju99 M gene consisted of mostly hydrophobic residues and contained three potential sites for asparagine (N)-linked glycosylation, two serine (S)-linked phosphorylation sites by protein kinase C, and two S- or threonine (T)-linked phosphorylation sites by casein kinase II.

• Reproductive and Developmental Biology
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• v.34 no.4
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• pp.283-288
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• 2010

During normal early embryonic development in mammals, the global pattern of genomic DNA methylation undergoes marked. changes. The level of methylation is high in male and female gametes. Thus, we cloned the cDNA of the porcine DNA methyltransferase 1 (Dnmt1) gene to promote the efficiency of the generation of porcine clones. In this study, porcine Dnmt1 cDNA was sequenced, and Dnmt1 mRNA expression was detected by reverse transcription-polymerase reaction (RT-PCR) in porcine tissues during embryonic development. The porcine Dnmt1 cDNA sequence showed more homology with that of bovine than human, mouse, and rat. The complete sequence of porcine Dnmt1 cDNA was 4,774-bp long and consisted of an open reading frame encoding a protein of 1611 amino acids. The amino acid sequence of porcine DNMT1 showed significant homology with those of bovine (91%), human (88%), rat (76%), and mouse (75%) Dnmt1. The expression of porcine Dnmt1 mRNA was detected during porcine embryogenesis. The mRNA was detected at stages of porcine preimplantation development (1-cell, 2-cell, 4-cell, 8-cell, morula, and blastocyst stages). It was also abundantly expressed in tissues (lung, ovary, kidney and somatic cells). Further investigations are necessary to understand the complex links between methyltransferase 1 and the transcriptional activity in cloned porcine tissues.

• Journal of Microbiology and Biotechnology
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• v.15 no.2
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• pp.321-329
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• 2005

Ornithine decarboxylase (ODC) antizyme is a key regulatory protein in the control of cellular polyamines. We have isolated two distinct ODC antizyme cDNA clones (AZS and AZL) from a flounder (Paralichthys olivaceus) brain cDNA library. Their sequences revealed that both clones required translational frameshifting for expression. Taking + 1 frameshifting into account, AZS and AZL products were 221 and 218 amino acid residues long, respectively, and shared $83.3\%$ amino acid sequence identity. Comparison of the structure and nucleotide sequence of the antizyme genes showed that the genes were highly conserved in flounder, zebrafish, mouse, and human. A phylogenetic tree was constructed, based on the antizyme amino acid sequences from various species. The presence of the two types of antizyme mRNA species in brain, kidney, liver, and embryo was confirmed by using the reverse transcription­polymerase chain reaction (RT-PCR) and Northern blot analysis. Recombinant proteins of flounder ODC antizymes, containing His-Nus-S tag at the amino-terminus, were overexpressed as His-AZL and His-AZS fusion proteins in Escherichia coli BL21 (DE3) pLys by using the pET­44a(+) expression vector.

• Food Science and Biotechnology
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• v.18 no.1
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• pp.207-212
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• 2009

Celandine (Chelidonium majus, family Papaveraceae) is an herb used extensively in traditional Korean medicine. To investigate its antioxidant and antiproliferative activities, the methanolic extract of celandine was introduced. The antioxidant properties of the extract were tested using various in vitro systems, including hydroxyl radical scavenging assay, DNA damage protection assay, 1,1-diphenyll-2-2-pricylhydrazyl (DPPH) free radical scavenging activity, metal chelating activity, and reducing power assay. The extract exhibited stronger antioxidant activity ($IC_{50}=7.92{\mu}g/mL$) against hydroxyl radicals in the Fenton system than butylated hydroxyanisole ($IC_{50}=51.46{\mu}g/mL$) and $\alpha$-tocopherol ($IC_{50}=67.48{\mu}g/mL$). Likewise, damage to the plasmid pBR 322 induced by hydroxyl radicals was found to be protected by the extract at a concentration of $400{\mu}g/mL$. Cellular proliferation and the induction of apoptosis were also examined by a cellular proliferation assay, flow cytometry, and mRNA expression analysis. Taken together, the extract significantly inhibited the growth of HT-29 cells in a concentration- and time-dependent manner, and gradually increased both the proportion of apoptotic cells and the expression of caspase-3. Overall, our research suggests that celandine possesses antioxidant and antiproliferative properties.

• Journal of Microbiology and Biotechnology
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• v.11 no.3
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• pp.529-533
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• 2001

The ${\beta}$-Amylase cDNA fragment from the oomcete Saprolegnia parasitica was cloned by reverse transcription-polymerase chain reaction (RT-PCR) using degenerate oligonucleotide primers derived from conserved ${\beta}$-amylase sequences. The 5'and 3'regions of the $\beta$-amylase gene were amplified using the rapid amplification of cDNA ends (rACE) system. It consisted of an open reading frame of 1,350 bp for a protein of 450 amino acids. Comparison between the genomic and cDNA sequences revealed that the intron was not present in the coding region. The deduced amino acid sequence of the ${\beta}$-amylase gene had a 97% similarity to the ${\beta}$-amylase of Saprolegnia ferax, followed by 41% similarity to those of Arabidopsis thaliana, Hordeum vulgare, and Zea mays. The ${\beta}$-amylase gene was also expressed in Saccharomyces cerevisiae by placing it under the control of the alcohol dehydrogenase gene (ADC1) promoter.

• The Journal of Korean Obstetrics and Gynecology
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• v.23 no.3
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• pp.78-90
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• 2010

Purpose: This study was performed to evaluate the effect of Clematidis Radix extract(CB) on osteoclast differentiation and gene expression. The osteocastogenesis and gene expression were determined in RANKL-induced RAW 264.7 cell. Methods: RANKL-induced RAW 264.7 cell with Clematidis Radix extract was stained by TRAP which is expressive marker of osteoclast. The gene expression of RANK, $TNF{\alpha}$, IL-6, iNOS and Cathepsin, those are factors related to bone resorption, was estimated by using Reverse Transcription-Polymerase Chain Reaction (RT-PCR). Results: Clematidis Radix extract decreased the number of TRAP-positive multi nuclei cell, and decreased the gene expression of RANK, $TNF{\alpha}$, IL-6, iNOS and Cathepsin K in RANKL-induced RAW 264.7 cell. Conclusion: It is concluded that Clematidis Radix extract might decrease the bone resorption resulted from decrease of osteoclast differentiation and it's related gene expression.