• Korean Journal of Veterinary Service
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• v.31 no.4
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• pp.485-492
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• 2008

Salmonella enterica serovars Gallinarum(SG, causative agent of fowl typhoid) and S Pullorum(SP, causative agent of pullorum disease) are very important bacterial pathogens in poultry industry. They share some common antigenic properties though the characteristics of outbreaks are quite different. To discriminate between SG and SP, we developed two rapid diagnostic methods, allele-specific real-time PCR and 3'-tailed PCR over 2 single nucleotide polymorphisms ($237^{th}\;and\;598^{th}$). In both methods, $237^{th}$ allele was found to be a good target for differential diagnosis, while $598^{th}$ allele produced some non-specific reactions.

• Journal of Animal Science and Technology
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• v.46 no.6
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• pp.897-902
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• 2004

To improve the methods used for the identification of Hanwoo, we performed a PCR using 3 -tailed primer associated with single nucleotide polymorphism(SNP) in Melanocortin 1 receptor(MCIR) gene. MCIR plays an important role in melanin synthesis and the SNP within MCIR was used as a target for PCRRFLP studies previously. A forward 3 -tailed primer, which matches with the template DNA of Hanwoo but not with others(blackhaired; Holstein and Black angus) at the site of 594th base sequence, and one reverse primer were designed for this study. When use this primer set, a size of 343bp was amplified by PCR only in Hanwoo, not in Holstein and Black angus. This result suggests that the PCR using our 3 -tailed primer would be very accurate, easy, reproducible and economic method to discriminate between Hanwoo meat and other black-haired ones.

• Korean Journal of Veterinary Service
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• v.33 no.4
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• pp.361-366
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• 2010

This study was carried out to investigate the characters of short-tailed dogs (DongGyeong dogs) with anatomical insights, molecular genetics in Gyeongju. The present study was conducted to further characterize of short-tailed dog population in Gyeongju. The short-tailed dog was analyzed in the distribution of 55 individual. The anatomical insights were by x-ray. For discovery of specific genes expressions were measured by Hot-start PCR analysis. Anatomy survey, the number of vertebral typical consists of more than 20. 88.9% of short-tailed dog populations consists of 3-8 vertebrates. The 54 individuals of the 47 observe the vestigial tailed of the sacrum. No detected sacrococcygeal vertebrae degradation individuals were malformation defects. The 3 genes were DEGs (differentially expressed genes) in Dong-Gyeong dogs. We succeeded in finding 3 novel DongGyeong dogs specific genes by using Hot-start PCR analysis, this study suggests that these novel genes may play role (s) in DongGyeong dogs.

• Korean Journal of Microbiology
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• v.37 no.4
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• pp.294-298
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• 2001

Highly effective polymerase chain reaction (PCR) often brings about false positivity caused by contamination of the sample with target nucleic acids. To solve this problem, in situ PCR (ISPCR) has been developed and applied onto various tissue sections and suspension cultures. With combination of PCR and in situ hybridization, this method amplifies the nucleic acid targets in situ and detect the amplified products inside the cells over the background of various cell types. In order to amplify the nucleic acid targets inside the cells, permeabilisation of a sample is required for the entry of amplification reactants into a cell. Treatments of a sample for the purpose allow not only the entry of reactants into the cell but also the exit of amplification products out of the cell. As a means to reduce the leakage of the amplification products, two methods were applied to suspension cultures of HIV-infected Molt/LAV and U 1.1 cells, in which modified, tailed primers produced long linear amplificants whereas biotinylated dUTP instead of dTTP did bulky products.

• BMB Reports
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• v.38 no.3
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• pp.350-353
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• 2005

Alanine at residue 73 (Ala-73) and aspartate at residue 9 (Asp-9) are characteristic to both Cw6 and Cw7 alleles of HLA-C gene and have been suggested as possible markers for psoriasis vulgaris (PsV). However, the results from various ethnic groups/populations are contradictory and inconclusive. In this study, an attempt has been made to examine the association between HLA-C (Ala-73 and Asp-9) and susceptibility to PsV among Saudi patients. Genomic DNA was extracted from 25 Saudi PsV patients and 75 control subjects. Polymerase chain reaction (PCR) was performed to amplify HLA-C sequences using earlier reported primers, C133P and C243PR. Sequence-specific primers were used to specifically detect nucleotide coding for Ala-73 and Asp-9 in all the subjects. The results showed significantly higher frequency of Asp-9 (84.0% versus 61.3%) in PsV patients as compared to controls (p < 0.05, 2-tailed Fisher's exact test). The frequencies of Ala-73 among PsV patients (92%) and controls (88%) did not differ significantly.

• Asian-Australasian Journal of Animal Sciences
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• v.29 no.3
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• pp.333-342
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• 2016

Lipins play dual function in lipid metabolism by serving as phosphatidate phosphatase and transcriptional co-regulators of gene expression. Mammalian lipin proteins consist of lipin1, lipin2, and lipin3 and are encoded by their respective genes Lpin1, Lpin2, and Lpin3. To date, most studies are concerned with Lpin1, only a few have addressed Lpin2 and Lpin3. Ontogenetic expression of Lpin2 and Lpin3 and their associations with traits would help to explore their molecular and physiological functions in sheep. In this study, 48 animals with an equal number of males and females each for both breeds of fat-tailed sheep such as Guangling Large Tailed (GLT) and Small Tailed Han (STH) were chosen to evaluate the ontogenetic expression of Lpin2 and Lpin3 from eight different tissues and months of age by quantitative real-time polymerase chain reaction (PCR). Associations between gene expression and slaughter and tail traits were also analyzed. The results showed that Lpin2 mRNA was highly expressed in perirenal and tail fats, and was also substantially expressed in liver, kidney, reproductive organs (testis and ovary), with the lowest levels in small intestine and femoral biceps. Lpin3 mRNA was prominently expressed in liver and small intestine, and was also expressed at high levels in kidney, perirenal and tail fats as well as reproductive organs (testis and ovary), with the lowest level in femoral biceps. Global expression of Lpin2 and Lpin3 in GLT both were significantly higher than those in STH. Spatiotemporal expression showed that the highest levels of Lpin2 expression occurred at 10 months of age in two breeds of sheep, with the lowest expression at 2 months of age in STH and at 8 months of age in GLT. The greatest levels of Lpin3 expression occurred at 4 months of age in STH and at 10 months of age in GLT, with the lowest expression at 12 months of age in STH and at 8 months of age in GLT. Breed and age significantly influenced the tissue expression patterns of Lpin2 and Lpin3, respectively, and sex significantly influenced the spatiotemporal expression patterns of Lpin3. Meanwhile, Lpin2 and Lpin3 mRNA expression both showed significant correlations with slaughter and tail traits, and the associations appear to be related with the ontogenetic expression as well as the potential functions of lipin2 and lipin3 in sheep.

• Molecules and Cells
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• v.26 no.3
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• pp.314-318
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• 2008

Korean long-tailed goral (Nemorhaedus caudatus) is one of the most endangered species in South Korea. However, detailed species distribution and sex ratio data on the elusive goral are still lacking due to difficulty of identification of the species and sex in the field. The primary aim of this study was to develop an economical PCR-RFLP method to identify species using invasive or non-invasive samples from five Korean ungulates: goral (N. caudatus), roe deer (Capreolus pygargus), feral goat (Capra hircus), water deer (Hydropotes inermis) and musk deer (Moschus moschiferus). The secondary aim was to find more efficient molecular sexing techniques that may be applied to invasive or non-invasive samples of ungulate species. We successfully utilized PCR-RFLP of partial mitochondrial cytochrome b gene (376 bp) for species identification, and sex-specific amplification of ZFX/Y and AMELX/Y genes for sexing. Three species (goral, goat and water deer) showed distinctive band patterns by using three restriction enzymes (Xbal, Stul or Sspl). Three different sexing primer sets (LGL331/335 for ZFX/Y gene; SE47/48 or SE47/53 for AMELX/Y gene) produced sex-specific band patterns in goral, goat and roe deer. Our results suggest that the molecular analyses of non-invasive samples might provide us with potential tools for the further genetic and ecological study of Korean goral and related species.

• Journal of Microbiology and Biotechnology
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• v.18 no.4
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• pp.624-630
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• 2008

Terminal restriction fragment length polymorphism (T-RFLP) is a method that has been frequently used to survey the microbial diversity of environmental samples and to monitor changes in microbial communities. T-RFLP is a highly sensitive and reproducible procedure that combines a PCR with a labeled primer, restriction digestion of the amplified DNA, and separation of the terminal restriction fragment (T-RF). The reliable identification of T-RF requires the information of nucleotide sequences as well as the size of T-RF. However, it is difficult to obtain the information of nucleotide sequences because the T-RFs are fragmented and lack a priming site of 3'-end for efficient cloning and sequence analysis. Here, we improved on the T-RFLP method in order to analyze the nucleotide sequences of the distinct T-RFs. The first method is to selectively amplify the portion of T-RF ligated with specific oligonucleotide adapters. In the second method, the termini of T-RFs were tailed with deoxynucleotides using terminal deoxynucleotidyl transferase (TdT) and amplified by a second round of PCR. The major T-RFs generated from reference strains and from T-RFLP profiles of activated sludge samples were efficiently isolated and identified by using two modified T-RFLP methods. These methods are less time consuming and labor-intensive when compared with other methods. The T-RFLP method using TdT has the advantages of being a simple process and having no limit of restriction enzymes. Our results suggest that these methods could be useful tools for the taxonomic interpretation of T-RFs.

• Korean Journal of Head & Neck Oncology
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• v.16 no.1
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• pp.14-19
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• 2000

Objectives: Epstein-Barr virus(EBV) is a B-lymphotrophic virus with a tumorigenic potential. EBV infection has been recognized as the main cause of nasopharyngeal carcinoma and Burkitt's lymphoma. Bcl-2 protein expression is known to be up-regulated by the EBV-latency associated antigen latent membrane protein(LMP). The aim of this study was to determine the incidence of EBV in squamous cell carcinomas of the larynx and the relationship between the presence of EBV and bcl-2 expression. Patients and Methods: From January 1994 to December 1977, 35 patients with primary squamous cell carcinoma of the larynx were studied. EBV genome DNA was surveyed by polymerase chain reaction(PCR) assay and then compared the results of in situ hybridization(ISH) for EBER1 using digoxigenin-tailed oligonucleotide probe. The expression of bcl-2 protein was studied by immunohistochemistry(IHC) using bcl-2 monoclonal antibody. Results: By PCR, EBV genome was detected in 22 of 35(62.9%) squamous cell carcinomas of the larynx. Nineteen of 35 cases(54.3%) showed a positive nuclear staining for EBER1 in tumor cells by ISH. Three cases showed positivity in inflammatory cells by ISH and one of them showed a positive staining of both tumor cells and inflammatory cells. Eighteen of 32 specimens(62.5%) were positive for bcl-2 protein. There was no significant correlations between the presence of EBV DNA and bcl-2 expression. Conclusions: It could be concluded that high incidence of EBV in the laryngeal cancer tissue may indicate a probable role of EBV in the development of laryngeal carcinoma.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.11
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• pp.4671-4676
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• 2014

Background: MicroRNA-200a (miR-200a) has been reported to regulate tumour progression in several tumours but little is known about its role in neuroblastoma. Our aim was to investigate the potential role and mechanism of miR-200a in neuroblastomas. Materials and Methods: Expression levels of miR-200a in tissues were determined using RT-PCR. The effect of miR-200a and shAP-$2{\gamma}$ on cell viability was evaluated using MTS assays, and target protein expression was determined using Western blotting and RT-PCR. Luciferase reporter plasmids were constructed to confirm direct targeting. Results were reported as mean${\pm}$S.E.M and differences were tested for significance using the 2-tailed Students t-test. Results: We determined that miR-200a expression was significantly lower in neuroblastoma tumors than the adjacent non-cancer tissue. Over-expression of miR-200 are reduced cell viability in neuroblastoma cells and inhibited tumor growth in mouse xenografts. We identified AP-$2{\gamma}$ as a novel target for miR-200a in neuroblastoma cells. Thus miR-200a targets the 3'UTR of AP-$2{\gamma}$ and inhibits its mRNA and protein expression. Furthermore, our result showed that shRNA knockdown of AP-$2{\gamma}$ in neuroblastoma cells results in significant inhibit of cell proliferation and tumor growth in vitro, supporting an oncogenic role of AP-$2{\gamma}$ in neuroblastoma. Conclusions: Our study revealed that miR-200a is a candidate tumor suppressor in neuroblastoma, through direct targeting of AP-$2{\gamma}$. These findings re-enforce the proposal of AP-$2{\gamma}$ as a therapeutic target in neuroblastoma.