• Title/Summary/Keyword: virus-free

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Strategies for Transmissible Gastroenteritis Virus(TGEV) Vaccine Production by Swine testicle cells

  • Kim, Min-Young;Jeong, Yeon-Ho;Kim, Myoung-Hwa;Ko, Yun-Mi
    • 한국생물공학회:학술대회논문집
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    • 2005.04a
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    • pp.203-206
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    • 2005
  • TGE (Transmissible Gastroenteritis) caused by a virus belonging to family coronavirus, results in an acute infection of the small intestine of the pig. The optimum operation variables such as multiplicity of infection (MOI), infection time and harvest time were investigated for TGE vaccine production by immobilized ST(swine testicle) cells. In the culture supplemented with 5% serum, maximum virus titer of $1.2{\times}10^6pfu/ml$ was obtained at the conditions of 0.01 MOI, 2day infection time, and 1 day harvest time. Serum is a potential source of bacterial, mycoplasmal and viral contamination, and it has a possibility of the introduction of serum proteins, prion and pyrogens into the final product. For these reasons, much attention has been focused on the development of serum-free media. A new serum-free media (SFM) has been developed in order to produce TGE vaccine of high quality with low cost. The performance of SFM developed was compared with other commercially available serum-free media and serum supplemented media in terms of virus productivity. The cultures with serum-free media showed higher titer than that with serum supplemented media. Among various serum-free media tested, CHO-S-SFMII showed highest virus titer.

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Molecular Analysis of double-stranded RNA in Abnormal Growing Oyster-Mushrooms, Pleurotos florida and P. ostreatus due to Virus Infection (Virus 이병(罹病) 느타리버섯 (Pleurotus)으로부터 double-stranded RNA 의 분리(分離))

  • Go, Seung-Joo;Park, Yong-Hwan;Shin, Gwan-Chull;Wessels, Josep G.H.
    • The Korean Journal of Mycology
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    • v.20 no.3
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    • pp.234-239
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    • 1992
  • The experiment was performed to find out the possibilities to detect virus infection in oyster mushrooms, Pleurotus species by analysis of doublestranded ribonucleic acid (ds RNA). Ds RNA segments were extracted from virus infected isolates which grew abnormally. But virus free isolates didn't show any ds RNA segments. The ds RNA was consisted of one large segment of 8100 base pairs (bp) and 4 smaller segments with 2170, 2120, 1980 and 1984 bp. Whereas, cell free virus particles showed only one larger ds RNA segment. The ds RNA was dissolved by RNase A in low salt, 0.1 M SSC and melted at $85^{\circ}C$. It was possible to use the ds RNA analysis for detecting virus infection directly from the host cells.

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Prevalence of Anti-bovine Leukemia Virus Antibodies in Dairy and Korean Native Cattle (우리 나라 젖소 및 한우의 Bovine Leukemia Virus 항체 분포조사)

  • 서국현;이정길;이채용;허태영;강석진;손동수;류일선;안병석;김남철
    • Journal of Veterinary Clinics
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    • v.20 no.2
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    • pp.172-176
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    • 2003
  • As a baseline study for the establishment of bovine leukemia virus(BLV)-free herd in Korea, the prevalence of anti-BLV antibody was determined in the present study. Sera from Korean native cows of 8 provinces and from dairy cattle of 9 provinces were subjected to enzyme-linked immunosorbent assay. Anti-BLV antibodies were positive in two (0.14%) of 1,413 Korean native cows. In contrast. 54.2% of 2,415 dairy cows were positive for anti-BLV antibodies, and their seropositive herd rate was 86.8%. And no differences were found in the sero-positive rates with age. The results indicate that the BLV infection rate has been increased continuously in Korea and that the establishment of BLV-free herd is imminent.

Viral Effects of a dsRNA Mycovirus (PoV-ASI2792) on the Vegetative Growth of the Edible Mushroom Pleurotus ostreatus

  • Song, Ha-Yeon;Choi, Hyo-Jin;Jeong, Hansaem;Choi, Dahye;Kim, Dae-Hyuk;Kim, Jung-Mi
    • Mycobiology
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    • v.44 no.4
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    • pp.283-290
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    • 2016
  • A double-stranded RNA (dsRNA) mycovirus was detected in malformed fruiting bodies of Pleurotus ostreatus strain ASI2792, one of bottle cultivated commercial strains of the edible oyster mushroom. The partial RNA-dependent RNA polymerase (RdRp) gene of the P. ostreatus ASI2792 mycovirus (PoV-ASI2792) was cloned, and a cDNA sequences alignment revealed that the sequence was identical to the RdRp gene of a known PoSV found in the P. ostreatus strain. To investigate the symptoms of PoV-ASI2792 infection by comparing the isogenic virus-free P. ostreatus strains with a virus-infected strain, isogenic virus-cured P. ostreatus strains were obtained by the mycelial fragmentation method for virus curing. The absence of virus was verified with gel electrophoresis after dsRNA-specific virus purification and Northern blot analysis using a partial RdRp cDNA of PoV-ASI2792. The growth rate and mycelial dry weight of virus-infected P. ostreatus strain with PoV-ASI2792 mycovirus were compared to those of three virus-free isogenic strains on 10 different media. The virus-cured strains showed distinctly higher mycelial growth rates and dry weights on all kinds of experimental culture media, with at least a 2.2-fold higher mycelial growth rate on mushroom complete media (MCM) and Hamada media, and a 2.7-fold higher mycelial dry weight on MCM and yeast-malt-glucose agar media than those of the virus-infected strain. These results suggest that the infection of PoV mycovirus has a deleterious effect on the vegetative growth of P. ostreatus.

Detection and Quantification of Apple Stem Grooving Virus in Micropropagated Apple Plantlets Using Reverse-Transcription Droplet Digital PCR

  • Kim, Sung-Woong;Lee, Hyo-Jeong;Cho, Kang Hee;Jeong, Rae-Dong
    • The Plant Pathology Journal
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    • v.38 no.4
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    • pp.417-422
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    • 2022
  • Apple stem grooving virus (ASGV) is a destructive viral pathogen of pome fruit trees that causes significant losses to fruit production worldwide. Obtaining ASGV-free propagation materials is essential to reduce economic losses, and accurate and sensitive detection methods to screen ASGV-free plantlets during in vitro propagation are urgently necessary. In this study, ASGV was sensitively and accurately quantified from in vitro propagated apple plantlets using a reverse transcription droplet digital polymerase chain reaction (RT-ddPCR) assay. The optimized RT-ddPCR assay was specific to other apple viruses, and was at least 10-times more sensitive than RT-real-time quantitative PCR assay. Furthermore, the optimized RT-ddPCR assay was validated for the detection and quantification of ASGV using micropropagated apple plantlet samples. This RT-ddPCR assay can be utilized for the accurate quantitative detection of ASGV infection in ASGV-free certification programs, and can thus contribute to the production of ASGV-free apple trees.

Development of a Reliable Technique to Eliminate Sweet potato leaf curl virus through Meristem Tip Culture Combined with Therapy of Infected Ipomoea Species

  • Cheong, Eun-Ju;Hurtt, Suzanne;Salih, Sarbagh;Li, Ruhui
    • Korean Journal of Plant Resources
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    • v.23 no.3
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    • pp.233-241
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    • 2010
  • In vitro elimination of Sweet potato leaf curl virus (SPLCV) from infected sweet potato is difficult due to low number of virus-free plants obtained from meristem tip culture and long growth period required for the virus detection. In this study, efficient production of the SPLCV-free sweet potato by in vitro therapy coupled with a PCR assay for virus detection was investigated. Infected shoots cultured on Murashige and Skoog medium were treated at three different temperatures for 7 weeks followed by meristem tip culture on the medium with or without ribavirin at 50 mg/L. The regenerated plantlets were tested for virus infection by a PCR assay. The results showed that the both heat- and cold-treatments, and addition of the ribavirin did not have significant effect on efficiency of the virus elimination. The meristem size, however, greatly affected the survival rate. Meristems sized over 0.4 mm survived better than smaller ones (0.2-0.3 mm). The PCR assay was approved to be a rapid, sensitive and reliable for the SPLCV detection in regenerated plantlets. Therefore, combination of cultivating meristem tips sized 0.4-0.5 mm on the medium at $22^{\circ}C$ without ribavirin and detection of SPLCV in the regenerated plantlets by the PCR assay was an efficient system for the SPLCV elimination from infected sweet potato.

Virus Free Stock Production by In vitro Stem Cutting of Shoot Tip Cultures of Grapes (포도 경정배양에서 얻은 유묘의 기내삽목에 의한 무병묘 생산)

  • 서정해;정재동;권오창
    • Korean Journal of Plant Tissue Culture
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    • v.28 no.2
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    • pp.81-85
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    • 2001
  • The experiment was conducted to know the effect of plant growth regulators on axillary bud elongation from in vitro stem cutting and the possibility of virus-free stock production. Axillary buds were well elongated in 3/4 strength MS medium supplemented with 0.1 or 0.5 mg/1 BA and 0.05 mg/1 NAA. Transferred plantlets could be established well in vermiculite and peat moss mixture (3:1, v/v) compare to other mixtures. In virus indexing, all the varieties of mother plants were infected by GLRV Ⅲ. Infected percentages of the three varieties were ranged from 30% to 75%. But negative response was revealed against the other species of virus, GLRV Ⅰ, GFLV and ArMV. Plantlet of 'Schuyler' and 'Muscat of Alexandria', which were cultured in vitro, showed positive response against GLRV Ⅲ and infected percentage of the former was 37.5% but the latter, 12.5%. On the other hand, that of 'Campbell Early' negativiely responded against all the species of virus indexed.

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Propagation and Attenuation of Japanese Encephalitis Virus in Tissue Culture Cells (조직배양세포에서의 일본뇌염virus 증식에 관한 연구)

  • Lee, Ho-Wang;Moon, Seok-Bae
    • The Journal of the Korean Society for Microbiology
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    • v.16 no.1
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    • pp.83-89
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    • 1981
  • Japanese encephalitis has been prevalent for long time in the Far East and many patients have been reported in both South East and Mid-West Asia recently. Recently, vaccine was used in prevention of this viral disease of man which was derived from formalin inactivated virus inoculated into mouse brain, but live attenuated active vaccine for human is not developed yet. Author inoculated Japanese encephalitis virus into several cell culture strains for development of live attenuated encephalitis virus strain and the results were as follows: 1. Japanese encephalitis virus was inactivated rapidly in cell free medium at $36^{\circ}C$ and totally inactivated by 72 hours. 2. In growth curve of Japanese encephalitis virus in HeLa cell cultures, maximal multiplication of the virus was occured at 4th day and virus multiplication was continued for at least 12 days. 3. After succeeding passage of the virus in HeLa cell cultures and human esophagus epithelial cell cultures, infectivity of virus for mice was disappeared from 2nd passage in HeLa cell cultures and 3rd passage in esophagus epithelial cell cultures. 4. In inoculation to monkey kidney epithelial cells and chick embryo cell cultures, infectivity of the virus for mice was continued after 10th passages.

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Studies on the Potato Virus X and Potato Leaf Roll Virus for Disease-free Seed Potato Production (무병종서 생산을 위한 감자X바이러스 및 엽권바이러스에 관한 연구)

  • Jhung-Il Choi
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.7 no.1
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    • pp.31-63
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    • 1969
  • A series of experiment was carried out to study on the production of disease-free seed potatoes at the Alpine Experiment Station from 1960 to 1968, which initiated a study of comparison on degeneration of plain warm region and high altitude products and the effect of latent potato virus X (PVX) and potato leaf roll virus(PLRV) on degeneration. Particular observations were made on some aspect of the nature of potato virus disease and its control such as concentrations of PVX, range of host plants, physical properties such as concentrations of PYX, range of host plants, physical properties and carrying effect of insects, by investigating 9 different areas of the main potato producing regions (Kimhae, Taegu, Choongju, Taejoen, Suwon, Kwangju, Chonju, Cheju and Chinju). Highly purified anti-serum was separated and tested for control of the virus disease and also various method of prevention and control of PLRV were observed, using cultivation of sprouted seed tubers, early harvesting method, and systemic chemicals. The results obtained are summarized as follows; 1. Potato yield in the plain region decreased by 32.8~66.3% in the first year cultivation of seed potatoes from colder region, and the rate of virus infection was 92.9 to 95.4%. 2. Plants of three families including, 20 species were susceptible to the PVX, and among the plants Salvia officinalis of a habits only was the carrier while the symptom of Digitalis purpurea of Screphulariaceae was masked. Necrosis and ring spot was occurred in most pJants of the Solanaceae and ring spot symptom also was observed in Nicotiana tabacum L. var. White Burley and in N. glutinosa. 3. The 8$C_2$ strain of virus had the following physical properties; thermal inactivation point, 68-$72^{\circ}C$ : dilution inactivation point, above 1, 000, 000 dilution: ageing in vitro, 240-360 days: and ageing in dry plant tissue, 30 days. 4. Myzus persicae and Oxya spp. did not transmit the 8$C_2$ strain of potato virus. 5. Virus was purified through the ammonium sulphate isolating method, and higher titer value, 1/2048 was obtained through anti-serum test. 6. Inhibition Chenopodiacae on the virus infection of potato was remarkable, and inhibition of local lesion host also was observed. 7, By earlier planting of sprouted seed tubers, growth period could be prolonged by 10 to 12 days. 8. Earlier harvest decreased much the rate of virus infection of seed potatoes. 9. According to the results of aphid control trial using systemic soil insecticides at Kangnung and Taekwanlyung, PSP 204, Disyston and Thimet was effective to aphid control. In particular, control effect of twice treatments of PSP 204 was great. 10. Treatmental effect of those chemicals lasted about 60-70 days. However, single foliar application of emulsified chemicals was not effective to potato virus control. 11. The effect of PSP 204, Disyston, and Thimet on the control of potato leaf roll virus was great, particularly in the case of two treatments of PSP 204, at Kangnung as well as at Taekwanlyung. Higher negative correlationship between the control effect of potato leaf roll virus and potato yield was observed showing the value r=-0.85 at Kangnung, and r=-0.87 at Taekwanlyung. 12. Differences in the control effects among PSP 204, Disyston, and Thimet was not noticed.

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Elimination of SPFMV from Virus-infected Sweet Potato Plants through Apical Meristem Culture

  • Kim, Young-Seon;Jeong, Jae-Hun;Park, Jong-Suk;Eun, Jong-Seon
    • Plant Resources
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    • v.7 no.3
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    • pp.200-205
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    • 2004
  • Sweet potato infected with a viral disease (SPFMV) showed irregular chlorotic patterns, so called feathering associated with faint or distinct ring spots that have purple-pigmented borders. SPFMV was eliminated from sweet potato plants using meristem tip culture. MS medium supplemented with BAP (2mg/L) and NAA (0.05 mg/L) was used for shoot proliferation and 1/2 MS medium for rooting of the plants. Highest percentage of regenerated plants (60%) was obtained from the optimum size (0.3-0.5mm) meristem tips. Of these, 60% plants were found negative for SPFMV by RT-PCR. Virus detection by RT-PCR was found to be a reliable method. Meristem-tip culture to produce SPFMV-free quality sweet potato and virus detection by RT-PCR is an efficient, time saving and reliable method for production of SPFMV-free tissue culture raised plants.

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