• KSBB Journal
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• v.9 no.3
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• pp.272-278
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• 1994

Attenuated varicella-zoster virus(VZV) was cultured in human embryonic lung cells. The effects of serum type and its concentration on the production of VZV were studied. Regardless of cell culture conditions, VZV yield was increased with multiplicity of infection, and the total cell concentration was decreased after virus infection. The newborn calf serum, calf serum, or horse serum was not as good as the fetal bovine serum or calf serum supplemented with iron for the propagation of VZV in the human embryonic lung cells. The yields of total VZV(cell-associated virus and cell-free virus) in the medium with calf serum containing iron were comparable to those in the medium supplemented with fetal bovine serum. Furthermore, some components of serum appear to be important for the maintenance of VZV infectivity.

• 한국생물공학회:학술대회논문집
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• 2002.04a
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• pp.271-272
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• 2002

Newcastle disease virus (NDV) vaccines were produced from Vero cells by using lively attenuated virus strain. The MOI of 0.1.' serum concentration of 2%. initial pH of 8.0. and infection time of 3 days were found to be optimum conditions for vaccine production. The treatment of polycation enhanced the virus production. When ascorbic acid was added as an antioxidant, NDV production was also enhanced. Utilization of $CaCl_2$ showed an inhibitory effect on the propagation of NDV. It was also found the ammonium ion concentration higher than 4mM inhibited virus production. Thus ammonium ion removal system was tried for the efficient production of NDV vaccine.

• Journal of Microbiology and Biotechnology
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• v.26 no.10
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• pp.1800-1807
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• 2016

To understand how human cytomegalovirus (HCMV) might change and evolve after reactivation, it is very important to understand how the nucleotide sequence of cultured HCMV changes after in vitro passaging in cell culture, and how these changes affect the genome of HCMV and the consequent variation in amino acid sequence. Strain JHC of HCMV was propagated in vitro for more than 40 passages and its biological and genetic changes were monitored. For each passage, real-time PCR was performed in order to determine the genome copy number, and a plaque assay was employed to get virus infection titers. The infectious virus titers gradually increased with passaging in cell culture, whereas the number of virus genome copies remained relatively unchanged. A linear correlation was observed between the passage number and the log₁₀ infectious virus titer per virus genome copy number. To understand the genetic basis underlying the increase in HCMV infectivity with increasing passage, the whole-genome DNA sequence of the high-passage strain was determined and compared with the genome sequence of the low-passage strain. Out of 100 mutations found in the high-passage strain, only two were located in an open reading frame. A G-T substitution in the RL13 gene resulted in a nonsense mutation and caused an early stop. A G-A substitution in the UL122 gene generated an S-F nonsynonymous mutation. The mutations in the RL13 and UL122 genes might be related to the increase in virus infectivity, although the role of the mutations found in noncoding regions could not be excluded.

• Journal of Microbiology and Biotechnology
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• v.29 no.12
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• pp.2006-2013
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• 2019

The isolation of respiratory viruses, especially from clinical specimens, often shows poor efficiency with classical cell culture methods. The lack of suitable methods to generate virus particles inhibits the development of diagnostic assays, treatments, and vaccines. We compared three inoculation methods, classical cell culture, the addition of a JAK2 inhibitor AZD1480, and centrifugation-enhanced inoculation (CEI), to replicate human respiratory syncytial virus (HRSV) and human metapneumovirus (HMPV). In addition, a combined method using AZD1480 treatment and CEI was used on throat swabs to verify that this method could increase virus isolation efficiency from human clinical specimens. Both CEI and AZD1480 treatment increased HRSV and HMPV genome replication. Also, the combined method using CEI and AZD1480 treatment enhanced virus proliferation synergistically. The combined method is particularly suited for the isolation of interferon-sensitive or slowly growing viruses from human clinical specimens.

• Research in Plant Disease
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• v.19 no.4
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• pp.288-293
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• 2013

In worldwide, viral diseases of apple plants has caused the serious problems like reduced production and malformation of fruits. Also, the damages of apple plants by virus and/or viroid infection (Apple chlorotic leaf spot virus, Apple stem grooving virus, Apple mosaic virus, and Apple scar skin viroid) were reported in Korea. However there is few report about the protection approach against the infection by apple viruses. Therefore, this paper introduced the experimental protocol for the development of virus-free apple cultivars (Danhong, Hongan, Saenara, Summerdream). Apple plants were treated at $37^{\circ}C$ for 4 weeks and shoot tips were cultured in vitro. After heat treatment, the detection of apple viruses was performed by RT-PCR using virusspecific detection primers in new apple cultivars. With the heat treatments followed by in vitro shoot tip culture, the proportion of virus-free stocks of 'Danhong', 'Hongan', 'Saenara', and 'Summerdream' was 28%, 16%, 12%, and 12%, respectively. Taken together, this approach can be a good tool for production of virus-free apple stocks.

• Journal of Microbiology and Biotechnology
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• v.11 no.2
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• pp.186-192
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• 2001

A primary culture of human fetal hepatocytes was obtained through a therapeutic abortion process at 26 weeks of gestation period. More than $10^8$ cells were seeded on a plastic plate. These hepatocytes were infected with hepatitis B virus (HBV). The HBV was purified from serum of one chronic HBV carrier. Transformed hepatocytes were subcultured in a 10% FBS-supplemented medium. The morphology of the transformed cell was epithelial-like. The cells from the first pass showed signs of early proliferation and had a latent period of more than 3 months after 6-7 passages. After the rest period, the transformed cell proliferated actively and they were subcultured every three days. Transformed hepatocytes were characterized by detection of the HBV transcript by RT-PCR. The secretion of virions from transformed cells was investigated by PCR with the cell medium. Two types of virions secreted into the culture medium were examined by using the transmission electron microscope. Another approach to study the secretion of virions in to culture medium was carried out with HBV antibody. HBsAg was detected in the culture medium of transformed cells using ELISA and Western blot analyses. These data suggested that the human fetal hepatocyte cell line has been established by infection of HBV, in which this cell line secreted viral particles into the culture medium.

• 한국생물공학회:학술대회논문집
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• 2000.11a
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• pp.615-618
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• 2000

The purpose of present study was to examine the efficacy of PAB (para-amino benzamidine) affinity column chromatography, pasteurization ($60^{\circ}C$ heat treatment for 10 h), and lyophilization steps, employed in the manufacture of urokinase from human urine, in the removal and/or inactivation of urine-born viruses. Bovine herpes virus (BHV) and Murine encephalomyocarditis virus (EMCV) were selected for this study. Samples from the relevant stages of the production process were spiked with the viruses and the amount of virus in each fraction was quantified by 50% tissue culture infectious dose ($TCID_{50}$). BHV and EMCV were effectively partitioned from urokinase during PAB chromatography with the log reduction factors of 6.71 and 5.27, respectively. Pasteurization was a robust and effective step in inactivating BHV and EMCV, of which titers were reduced from initial titers of $8.65\;log_{10}\;TCID_{50}$ and $7.81\;log_{10}\;TCID_{50}$, respectively, to undetectable levels within 1 hour of treatment. The log reduction factors achieved during lyophilization were 2.06 for BHV and 4.54 for EMCV. These results indicate that the production process for urokinase has sufficient virus reducing capacity to achieve a high margin of virus safety.

• Korean Journal of Veterinary Service
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• v.32 no.2
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• pp.103-109
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• 2009

Influeza type A virus have been worldwide problematic in animals as well as in humans. In this study, the use of reverse-transcriptase polymerase chain reaction (RT-PCR) was described for detecting influenza virus type A. The primer of RT-PCR was designed from an nonstructural (NS) gene of Influenza A virus. By RT-PCR, a product with the size of 189 bp was detected only when influenza virus type A was used as template. No products could be detected with Influenza virus type B as well as other respiratory pathogens. The detection limit of the RT-PCR was up to $10^{0.3}TCID_{50}$ which is comparable to the sensitivity of cell culture method. The RT-PCR could detect the influenza A virus from nasal turbinates of the ferrets infected with influenza virus type A not type B.

• Korean Journal Plant Pathology
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• v.12 no.3
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• pp.358-362
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• 1996

감자 잎말림 바이러스를 검정하기 위하여 ELISA 및 전자현미경에 의해 바이러스 감염이 확인된 기내 배양중인 감장의 줄기로부터 RT-PCR 분석을 수행하였다. 분리된 총 RNA들로부터 바이러스 cDNA를 합성하고 감자 잎말림 바이러스 외피단백질의 일부인 465bp를 특이하게 증폭하도록 고안한 두 primer를 사용하여 PCR 반응을 하였다. 증폭된 465pb의 DNA 절편의 염기서열을 분석한 결과 역시 감자 잎말림 바이러스임을 확인하였다. 바이러스 검정에 있어서 EL-ISA 방법과 RT-PCR 방법간의 민감도를 조사한 결과 RT-PCR 방법간의 민감도를 조사한 결과 RT-PCR 방법이 ELISA 방법보다 감자 잎말림 바이러스검정에 있어서보다 정확한 방법인 것으로 사료된다.

• Journal of the Korea Institute of Information and Communication Engineering
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• v.21 no.6
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• pp.1212-1217
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• 2017

We constructed a population of myelinated cells with co-culture of neuronal cells and Schwann cells from DRG. Schwann cells and neuronal cells were isolated from dorsal root ganglion (DRG) in embryos of rat in vitro respectively. The cultured Schwann cells and cultured neuronal cells, respectively were co-cultured in a same plate. This procedure contains following four steps: first step of suspension of the embryonic dorsal root ganglion cells, second step of addition of anti-mitoticcocktail, third step of purification of dorsal root cells, and fourth step of addition of Schwann cells to dorsal root ganglion cells. These cells were performed accomplishment of myelination. This myelinated co-culture system was infected by Semliki forest virus and then induced demyelination processing in this myelinated co-culture. We identified myelination and demyelination processing using antibody of peripheral myelin protein 22 (PMP 22) meaning presence of myelinated neuron.