• Korean Journal of Microbiology
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• v.47 no.2
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• pp.117-123
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• 2011

The purpose of this study was to evaluate and compare different elution and concentration methods for optimization of human rotavirus (HRV) detection method using real-time RT-PCR and cell culture techniques. The leafy vegetable samples (lettuce, Chinese cabbage) were artificially inoculated with HRV. Viruses were extracted from the vegetables by two different elution buffers, buffer A (100 mM Tris-HCl, 50 mM glycine, 3% beef extract, pH 9.5) and buffer B (250 mM Threonine, 300 mM NaCl, pH 9.5), and the extracted viruses were concentrated by filtration and PEG precipitation sequentially. To determine infectivity of the viruses, the viruses recovered from the samples were infected to the MA-104 cells, and integrated cell culture real-time RT-PCR was performed at 1, 48, 72, 96, 120, 144, 168 h post-infection (p.i.). The elution buffer A was more efficient in extracting the virus from the produce samples tested than the buffer B, 29.54% and 18.32% of recoveries, respectively. The sensitivity of real-time RT-PCR method was markedly improved when the virus was concentrated by the filtration method. When the viruses were eluted and concentrated by buffer A and filtration, respectively, the average recovery rate was approximately 51.89%. When the viruses recovered from samples were infected to MA-104 cell, infectious HRV was detected within 48 h p.i. by ICC/real-time RT-PCR, whereas cytopathic effects were not observed until 72 h p.i. The optimized detection method evaluated in this study could be useful for rapid and reliable detection of HRV in fresh produce products and applied for detection of other food-borne viruses.

• Development and Reproduction
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• v.11 no.3
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• pp.253-261
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• 2007

Lactadherin is a glycoprotein of human milk fat globule membrane that binds to mucin and butyrophilin forming the protein complex. Especially, mucin and lactadherin in human milk efficiently protect infants with poor immune functions right after birth from infections by microorganisms and play important roles for their early survival, growth and development. Lactadherin inhibits the propagation and growth of rotavirus that is a global pathogen causing infants' diarrhea. Recently this protein was known to promote neovascularization and its deficiency related to develop Alzheimer's disease. In this study, the basic biochemical and physiological aspects of lactadherin were investigated. Messenger RNAs were isolated from mammary tissues from Korean women patients to clone a 1.2 kb cDNA and sequenced its DNA to determine its amino acid sequences. The cDNA was cloned to express its 43 kD protein in E. coli, which was confirmed by Western blot. The recombinant protein was purified and injected to 2 rabbits to raise antibodies against it. The semi-purified milk fat globule membrane proteins from Korean women was analyzed by Western blot using the rabbit antibody to give 70, 55, 46, 30 kD bands. Also several polymorphism and SNPs of lactadherin gene from Korean women were observed compared with those of Caucasian women.

• Pediatric Gastroenterology, Hepatology & Nutrition
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• v.4 no.1
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• pp.34-40
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• 2001

Purpose: The purpose of this study was to evaluate the clinical efficacy of reverse transcription-polymerase chain reaction (RT-PCR) in detecting rotaviral gastroenteritis in children comparing with that of commercial immunoassays. Methods: Stools from 79 children admitted Korea University Hospital due to diarrhea were collected from December 1999 to February 2000. Immunoassays were done using commercial rotavirus Latex kit and Rotatec (ELISA) kit. RT-PCR was performed to amplify group A rotavirus, most commonly pathogenic to human, using VP4- and VP7-specific primers. The detection rates of immunoassays and RT-PCR were compared. Results: ELISA assay was superior to LA assay and moderately concordant with RT-PCR in detecting rotaviral gastroenteritis. Conclusion: Although RT-PCR is known very sensitive, it does not have significant advantage over immunoassay in detecting rotaviral gastroenteritis.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.93-93
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• 2003

모유의 뮤신 복합체는 rotavirus에 특이적으로 결합하여 항 바이러스활동을 보여주는 것으로 나타났다. 자연 상태에서의 뮤신은 몇몇 작은 분자들과 복합적으로 연합되어 있는데 70kda의 glycoprotein, butyrophilin, 그리고 glycosylated component, lactadherin을 포함하고 있다. 그중 rotavirus에 가장 높은 결합력과 항바이러스 활동을 나타내는 Lactadherin은 모유의 유단백질의 하나인 뮤신과 결합되어 분비되는 당단백질의 하나로 분자량이 46kda이고, 지방구막 속에 연합되어있다. 이러한 배경에서 본 연구에서는 한국 여성의 breast tissue로부터 lactadherin 유전자의 cloning 및 in vitro에서의 발현을 유도하여 lactadherin band만을 purify하였고 여기서 얻은 lactadherin을 항체 생산을 위한 항원으로 사용하여 anti-lactadherin antibody를 확보하였다. 이 항체는 human milk에서의 lactadherin을 동정하는데 사용하였는데 western blot결과 lactadherin을 포함하여 몇 몇 단백질들이 확인되었다. Human milk내 mucin은 몇몇 작은 분자들과 복합체를 형성하는 것으로 확인되어졌는데 70kda의 glycoprotein, butyrophilin 그리고 46kda의 glyxosylated component, lactadherin을 포함하고 있는 milk mucin은 associated molecular임을 확인할 수 있었다.

• Journal of Ginseng Research
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• v.40 no.4
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• pp.309-314
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• 2016

Korean Red Ginseng (KRG) is a heat-processed ginseng developed by the repeated steaming and air-drying of fresh ginseng. Compared with fresh ginseng, KRG has been shown to possess greater pharmacological activities and stability because of changes that occur in its chemical constituents during the steaming process. In addition to anticancer, anti-inflammatory, and immune-modulatory activities, KRG and its purified components have also been shown to possess protective effects against microbial infections. Here, we summarize the current knowledge on the properties of KRG and its components on infections with human pathogenic viruses such as respiratory syncytial virus, rhinovirus, influenza virus, human immunodeficiency virus, human herpes virus, hepatitis virus, norovirus, rotavirus, enterovirus, and coxsackievirus. Additionally, the therapeutic potential of KRG as an antiviral and vaccine adjuvant is discussed.

• Pediatric Infection and Vaccine
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• v.22 no.2
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• pp.81-90
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• 2015

Purpose: Group A rotavirus (RV) is most common etiologic agent of acute gastroenteritis (AGE) in children worldwide. Recently, vaccination has been introduced in several countries to reduce the disease burden caused by RV infections, but continuous surveillance of RV strains is necessary to detect the emergence of potential variants induced by vaccine-immune pressure. This study aimed to investigate the changing pattern of RV genotypes in children with AGE, following the introduction of vaccination in Korea. Methods: Genotyping of RVs by RT-PCR on the basis of VP7 and VP4 gene segment sequence was carried out on 201 rotavirus-positive stool samples, from children hospitalized with AGE between August 2009 and June 2013. We have directly sequenced PCR products and analyzed the phylogenetic tree. Results: The most prevalent G genotype was G9 (33.3%), followed by G1 (22.4%), G3 (15.9%), G2 (6.0%), G4 (3.0%), G10 (1.5%), and mixed G-type (15.4%), with some nontypeable cases (2.5%). The detected P genotypes were P[4] (45.3%), P[8] (43.8%), mixed P-type (10.4 %), and P[2] (0.5%). The G9P[4] genotype was predominantly observed in hospitalized cases in Seoul in 2010/2011, however G1P[8] has been re-emerged as the predominant genotype in the following season (P =0.004). Conclusions: It seems that the periodic fluctuation in predominance of the G1, G3, and G9 strains occurred in Korea during 2009-2013, following the introduction of RV vaccination.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2002.11a
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• pp.94-94
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• 2002

Lactadherin (formerly known as BA46), a major glycoprotein of the human milk fat globule membrane, is abundant in human breast milk and breast carcinomas and may prevent symptomatic rotavirus infections. In this study, under the control of mouse whey acidic protein (WAP) promoter, the expression pattern of lactadherin (Ltd) in lactogenic hormone-dependent mouse mammary epithelial cell line HC11 were tested. pLNWLtd construct containing 2.4 kilobases of the WAP promoter and 1.5 kilobases of human lactadherin gene was stably transfered into HC11 cells using retroviral vector system. Integration and expression level of the transgene was estimated using PCR and RT-PCR, respectively. Prominent induction of Ltd gene under the WAS promoter was accomplished in the presence of insulin, hydrocortisone and prolactin, while induction with insulin alone resulted in lower expression. Our results demonstrate that the expression of the transgene is increased by synergistic effect of several lactogenic hormones, including insulin, hydrocortisone, and prolactin.

• The Journal of the Korean Society for Microbiology
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• v.34 no.5
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• pp.453-459
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• 1999

Astrovirus is frequently associated with diarrhea in children. It can not be readily isolated by cell culture, and an electronmicroscope is usually used for detection of this agent. Recently in 1995 a combined method of reverse transcription-polymerase chain reaction (RT-PCR) was designed for easier detection of astrovirus, which is based on the conserved sequence in 3'-end of genomes of the 7 known serotypes of human astrovirus. As of yet there has not been any report of astrovirus data in Korea using the RT-PCR methods. The purpose of this study was to detect astrovirus incidence, severity of symptoms, seasonal variation and co infection rate with rotavirus in Korean children inpatients with diarrhea. Fecal specimens from 61 young children hospitalized with gasteroenteritis Korea from Jan. 1996 through Mar. 1997. They were examined for astroviurs infection by RT-PCR method. Results are as follows:1. Astrovirus was detected at 9.8% (6/61) from fecal specimens of children with severe diarrhea by EIA using monoclonal antibody coated plates. 2. Astorvirus was detected at 29.5% (18/61) from fecal specimens of children with severe diarrhea by RT-PCR. 3. The age of the 18 children affected by astrovirus ranged from 2 monthes to 7 years with mean of 3.0 years. 4. Mean hospital stay of the 18 children was 6.1 days. 5. Five (27.8%) astrovirus RT-PCR positive strains were confirmed in November and in December, respectively out of 18 specimens in total. 6. Astrovirus coinfection with rotavirus type G1 was confirmed in 15/16 specimens (93.8%), and with type G2 was in 1/16 specimens (6.3%).

• The Journal of Korean Society of Virology
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• v.28 no.3
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• pp.247-265
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• 1998

To determine the sequence and expression of the VP7 gene of Korean isolates (CAU-9), viral RNA was purified and used for cDNA amplification by RT-PCR. The VP7 cDNA was cloned, sequenced, and expressed using baculovirus expression system. The result showed that the sequence homologies CAU-9 compared with foreign isolated strains Wa, 417, TMC-II, 95B and SA11 were ranged from 74.0% to 95.1 % of nucleotide sequence and 35% to 43% of amino acid sequence, respectively. High homology of CAU-9 was observed in Japanease isolates 417 (nucleotide sequence homology was 95.1% and amino acid sequence homology was 43%). To express VP7 gene, the VP7 cDNA was cloned into pCR-Bac vector and inserted into the genome of baculovirus adjacent to the polyhedrin promoter by cotransfection of Spodoptera frugiperda (Sf9) insect cells with wild type baculovirus DNA. In antigenic analysis of Sf9 cells inoculated with the recombinant VP7, immunofluorescence assay revealed positive for viral antigens. In metabolic labeling of Sf9 cell lysates infected with recombinant baculoviruses, it was revealed that the protein of 34 kDa was expressed. The limited study of expressed VP7 protein inoculated with guinea pigs failed to elicit neutalizing antibody. As a results, the sequence analysis and expression of VP7 protein of rotavirus CAU-9 isolated from an infant in Korea could permit the conformation and development of virus like particles which may be useful in designing vaccine strategy.

• Korean Journal of Animal Reproduction
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• v.27 no.1
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• pp.15-23
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• 2003

Lactadherin (formerly known as BA46), a major glycoprotein of the human milk fat globule membrane, is abundant in human breast milk and breast carcinomas and may prevent symptomatic rotavirus infections. In this study, under the control of tissue specific and hormonal inducible mouse whey acidic protein (WAP) promote., the expression pattern of lactadherin (Ltd) in lactogenic hormone-dependent mouse mammary epithelial cell line HC11 were tested. pLNWLtd construct containing 2.4 kilobases of the WAP promoter and 1.5 kilobases of human lactadherin gene was stably transfered into HC11 cells using retroviral vector system. Integration and expression level of the transgene was estimated using PCR and RT-PCR, respectively. Prominent induction of Ltd gene under the WAP promoter was accomplished in the presence of insulin, hydrocortisone and prolactin. Compared to the control (cells cultured with insulin alone), however we observed that the WAP promoter was leaky. These data indicate that luther studies are needed in finding an appropriate promoter other than WAP promoter because of its leakiness.