• Journal of Ginseng Research
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• v.21 no.3
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• pp.147-152
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• 1997

Ginsenosides $Rh_1$ and $Rh_2$ Induced the differentiation of F9 teratocarcinoma stem cells. These agents are structurally similar to the steroid hormones, therefore, we speculated that the steroid receptor (s) or novel nuclear receptor (s) could be involved in the differentiation process induces by them. Based on this speculation, we tried to alone new nuclear receptors with reverse transcription-polymerase chain reaction (RT-PCR) method by isolating RNA from F9 teratocarcinoma cells induced by ginsenosides. By using RT-PCR with degenerated primers from highly conserved DNA binding domain of nuclear receptors, we identified several nuclear receptors. In northern blot analysis we found that these clones are transcriptionally regulated by ginsenoside Rhl or Rh2 treatment. Further characterizations of these clones are needed to identify the mechanism of gene expression, which has an important role in the differentiation of F9 cells induced by ginsenosides.

• Journal of Microbiology and Biotechnology
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• v.15 no.4
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• pp.838-843
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• 2005

Brain-derived neurotrophic factor (BDNF) is a small secretory protein and a member of the nerve growth factor (NGF) gene family. We cloned the flounder BDNF gene from a flounder brain cDNA library. The nucleotide sequence of the cloned gene showed an open reading frame (ORF) consisting of 810 bp, corresponding to 269 amino acid residues. The tissue distribution of flounder BDNF was determined by reverse transcription-polymerase chain reaction (RT-PCR) in brain, embryo, and muscle tissues. To express fBDNF using a eukaryotic expression system, we constructed the vector mpCTV-BDNF containing the fBDNF gene and transformed this vector into Chlorella ellipsoidea. Stable integration of introduced DNA was confirmed by PCR analysis of genomic DNA, and mRNA expression in C. ellipsoidae was confirmed by RT-PCR analysis.

• Research in Plant Disease
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• v.7 no.1
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• pp.1-7
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• 2001

An isolate of Cucumber mosaic cucumovirus(CMV) was isolated from Hydrangea macrophylla for. otaksa(Sieb. et Zucc. ) Wils. showing mosaic symptoms, and designated as Hm-CMV. Hm-CMV was characterized by the tests of host range, physical properties, serological properties, RNA and coat protein compositions, and reverse transcription and polymerase chain reaction (RT-PCR) analysis. Twelve species in 4 families were used in the host range test of Hm-CMV and could be differentiated from Y-CMV used as a control CMV by the ringspot and line pattern on inoculated leaves of several tobacco plants. Thevirus produced local lesions on inoculated leaves of Chenopodium amarticolor, C. quinoa and Vigna unguiculata. The physical properties of the virus were as follows; thermal inactivation point(TIP) was 60$\^{C}$, dilution end point (DEP) was 10$\^$-3/, and longevity in vitro (LIP) was 3∼4 days. Hm-CMV was serologically identical to Y-CMV. SDS-polyaciylamide gel electrophoresis(SDS-PAGE) showed one major protein band of about 28 kDa. In RNA or dsRNA analysis, Hm-CMV consisted of four RNA or dsRNA species, but satellite RNA was not detected. In RT-PCR using CMV-common primer and CMV subgroup I-specific primer, bothe amplified expected size of about 490 bp and 200 bp DNA fragments from Hm-CMV, respectively. Restriction enzyme analysis of the 490 bp RT-PCR products using EcoR I and Msp I showed that Hm-CMV belonged to CMV subgroup I. However, Hm-CMV could be differentiated from other CMV subgroup I isolates by RNA fingerprinting by arbitrarily primed polymerase chain reaction (RAP-PCR).

• The Journal of Korean Medicine
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• v.23 no.2
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• pp.57-69
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• 2002

Objective: The aim of this study is to investigate the effect of Injin butanol fractions with Thin Layer Chromatography on Fas-mediated Apoptosis. Method: Injin-butanol fraction separated by TLC. MIT assay, cell cycle analysis, Caspase-3 protease assay, DNA fragmentation assay and quantitative RT-PCR were performed to evaluate the effects of TLC extraction of lnjin-butanol fraction on cell viability, cell cycle progression and apoptosis. Results: Scopoletin, luteolin, apigenin and unknown powder was isolated by TLC. Fas-mediated apoptosis analysis shows that scopoletin has inhibiting function on apoptosis. Caspase- 3 protease assay analysis shows that scopoletin inhibits activity of caspase-3. Quantitative RT-PCR analysis shows that no activity on caspase-3, but apoptosis inhibition cytokine -Bcl-2- is activated, and apoptosis activating cytokine -Bax- is unactivated. Conclusion: These results show that each fraction of Injin-butanol TLC extraction, especially scopoletin, acts as a protective function on liver cell viability, and inhibitory function on apoptosis. (J Korean Oriental Moo 2002;23(2):57-69)

• Korean Journal of Veterinary Research
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• v.36 no.3
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• pp.627-633
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• 1996

To analyze the genomic diversity of transmissible gastroenteritis virus (TGEV), the N-terminal half of the spike (S) glycoprotein gene and nonstructural protein gene (open reading frames 3 and 3-1) were amplified by reverse transcriptase reaction and polymerase chain reaction (RT-PCR), and analyzed using restriction fragment length polymorphism (RFLP) patterns of the amplified DNA. In this study, TGEV Miller (M6) and Purdue (P115) strains were used as reference strains, and two vaccine strains (MSV and STC3) and four Korea isolates (P44, VRI-WP, VRI-41, and VRI-48) were analyzed. All TGEV strains were amplified with three TGEV primer pairs. Although there was some exception in RFLP analysis, this method differentiated TGEV strains into following groups : Miller group (M6 and MSV), Purdue group (PUS, STC3, P44, VRI-WP, VRI-41, and VRI-48). Using Sau3AI and SspI, VRI-48 was differentiated from the Miller and Purdue type viruses. The RT/PCR in conjuction with RFLP analysis was a rapid and valuable tool for differentiating several strains of TGEV. This study revealed the occurences of distinct difference in genome of TGEV strains.

• Korean Journal of Veterinary Research
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• v.39 no.1
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• pp.104-110
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• 1999

Field infectious bursal disease viruses (IBDVs) were isolated from IBDV-suspected commercial chickens. The variable region in VP2 gene of six Korean IBDV isolates (K-IBDVs) and IBD vaccines was examined using the reverse transcriptase / polymerase chain reaction-restriction fragment length polymorphism (RT/PCR-RFLP) assay. With all K-IBDVs and vaccine IBDVs, a 474-bp fragment of the VP2 gene was amplified and tested with various restriction enzymes. Restriction enzymes BstNI and StyI differentiated K-IBDV isolates and IBD vaccines into four groups. Restriction enzyme profiles of K-IBDV isolates were different from them of IBD vaccines. K-IBDV isolates except for 310 isolate had specific SspI and TaqI recognition sites, which were recognized in highly virulent IBDVs, but IBD vaccines had no those sites. This study showed that RT/PCR-RFLP assay was thought to be valuable tool for differentiation of IBDVs and identification of highly virulent IBDV.

• Korean Journal of Breeding Science
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• v.43 no.5
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• pp.448-456
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• 2011

We used suppression subtractive hybridization (SSH) combined with mirror orientation selection (MOS) method to screen differentially expressed genes from red-fleshed kiwifruit 'Hongyang'. As a result, the 288 clones were obtained by subcloning PCR product and 192 clones that showed positive clones on colony PCR analysis were selected. All the positive clones were sequenced. After comparisons with the NCBI/Genbank database using the BLAST search revealed that 30 clones showed sequence similarity to genes from other organisms; 10 clones showed significant sequence similarity to known genes. Among these clones, 3 clones (AcF21, AcF42 and AcF106) had sequence homology to 1-aminicyclopropane-carboxylic acid (ACC)-oxidase (ACO) that known to be related to fruit ripening. The expression patterns of differentially expressed genes were further investigated to validate the SSH data by reverse transcription PCR (RT-PCR) and quantitative real-time PCR (qReal-time PCR) analysis. All the data from qReal-time PCR analysis coincide with the results obtained from RT-PCR analysis. Three clones were expressed at higher levels in 'Hongyang' than 'Hayward'. AcF21 was highly expressed in the other genes at 120 days after full bloom (DAFB) and 160 DAFB of 'Hongyang'.

• Research in Plant Disease
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• v.16 no.3
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• pp.247-253
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• 2010

Apple scar skin viroid (ASSVd) is one of the smallest viral pathogens infecting fruits, especially apple, and causes a significant damage to fruit trees. ASSVd usually induced the skin-dapple ring symptoms, but in 'Fuji' varieties, corked spot were occurred on the fruit skin in 2009. This new symptom will be of great helpful to diagnosis ASSVd in sight. ASSVd was surveyed in apple and pear from 2009 to 2010 in Korea, and ASSVd was identified in 20 out of 1,193 trees. The infection rate was 1.7%. To screen the infectivity of ASSVd among apple cultivars, real-time RT-PCR was applied followed by designing of ASSVd specific primers based on highly conserved regions of several ASSVd isolates including Korean isolate. NADH dehydrogenase subunit 5 (nad 5) gene, which is mRNA of the mitochondrial gene, was used for internal control. In this study, ASSVd infected apples were classified into 12 groups depending on different symptoms and symptom severity (scaring, rusting or malformation). Taken together, this study suggested that real-time PCR analysis was more sensitive to detect the low copy of ASSVd on early viroid infected apple skins than regular RT-PCR method.

• Journal of Veterinary Science
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• v.23 no.6
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• pp.80.1-80.6
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• 2022

A 10-year-old male neutered Labrador Retriever presented with a history of acute-onset tachypnoea, lethargy and anorexia. The dog was pyrexic, tachypnoeic and dyspnoeic on examination. A rapid antigen test for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) was performed on an oropharyngeal swab and yielded a positive result. SARS-CoV-2 infection was subsequently confirmed by reverse transcription polymerase chain reaction (RT-PCR) analysis. Both of the dog's owners had positive rapid antigen test and RT-PCR analysis results for SARS-CoV-2. Additional diagnostics included computed tomography. Resolution of the dog's clinical signs was achieved with symptomatic treatment.

• The Plant Pathology Journal
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• v.27 no.4
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• pp.301-309
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• 2011

The filamentous fungus Fusarium graminearum is an important cereal pathogen. Although quantitative realtime PCR (qRT-PCR) is commonly used to analyze the expression of important fungal genes, no detailed validation of reference genes for the normalization of qRT-PCR data has been performed in this fungus. Here, we evaluated 15 candidate genes as references, including those previously described as housekeeping genes and those selected from the whole transcriptome sequencing data. By a combination of three statistical algorithms (BestKeeper, geNorm, and NormFinder), the variation in the expression of these genes was assessed under different culture conditions that favored mycelial growth, sexual development, and trichothecene mycotoxin production. When favoring mycelial growth, GzFLO and GzUBH expression were most stable in complete medium. Both EF1A and GzRPS16 expression were relatively stable under all conditions on carrot agar, including mycelial growth and the subsequent perithecial induction stage. These two genes were also most stable during trichothecene production. For the combined data set, GzUBH and EF1A were selected as the most stable. Thus, these genes are suitable reference genes for accurate normalization of qRT-PCR data for gene expression analyses of F. graminearum and other related fungi.