• Reproductive and Developmental Biology
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• v.31 no.4
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• pp.267-272
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• 2007

Human embryonic stem (ES) cells are derived from the inner cell mass of the preimplantation embryo and have the capacity to differentiate into various types of cells in the body. Hence, these cells may potentially be an indefinite source of cells for cell therapy in various degenerative diseases including neuronal disorders. For clinical applications of human ES cells, directed differentiation of these cells would be necessary. The objective of this study is to develop the culture condition for the expansion of neural precursor cells derived from human ES cells. Human ES cells were able to differentiate into neural precursor cells upon a stepwise culture condition. Neural precursor cells were propagated up to 5000-fold in cell numbers over 12-week period of culture and evaluated for their characteristics. Expressions of sox1 and pax6 transcripts were dramatically up-regulated along the differentiation stages by RT-PCR analysis. In contrast, expressions of oct4 and nanog transcripts were completely disappeared in neural precursor cells. Expressions of nestin, pax6 and sox1 were also confirmed in neural precursor cells by immunocytochemical analysis. Upon differentiation, the expanded neural precursor cells differentiated into neurons, astrocytes, and oligodendrocytes. In immunocytochemical analysis, expressions of type III ${\beta}$-tubulin and MAP2ab were observed Presence of astrocytes and oligodendrocytes were also confirmed by expressions of GFAP and O4, respectively. Results of this study demonstrate the feasibility of long-term expansion of human ES cell-derived neural precursor cells in vitro, which can be a potential source of the cells for the treatment of neurodegenerative disorders.

• Research in Plant Disease
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• v.13 no.3
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• pp.177-182
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• 2007

In this experiment, 236 isolates of Botrytis cinerea isolated from the lesions of ginseng grey mold in 2005 and 2006 were examined for their sensitivity to fungicide inhibiting ${\beta}-tubulin$ assembly. The baselines of fungicide resistance were determined as 10.0 and $0.2{\mu}g/ml$ of $EC_{50}$ values for carbendazim and the mixture of carbendazim and diethofencarb, respectively. The ratios of isolates resistant to carbendazim in 2005 and 2006 was investigated to be 87.6 and 96.6%, respectively. In the case of the mixture of carbendazim and diethofencarb, the ratio of the resistant isolates was 23.6% for 2005 and 24.5% for 2006. The ratio of the resistant isolates to the mixed fungicide was fluctuated according to regions where isolates of B. cinerea were obtained. In Yeoncheon of Gyeonggi Province, 4.3% of the isolates used in the experiment was resistant in 2005 and no resistant isolate was obtained in 2006. Among 5 regions, the ratio of resistant isolates was the highest as 70.0% in Yecheon of Gyeongbuk Province.

• Clinical and Experimental Reproductive Medicine
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• v.28 no.1
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• pp.33-40
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• 2001

Objective: This study was to establish the human embryonic stem (ES) cells derived from frozen-thawed blastocyst stage embryo that were destined to be discarded after five years in routine human IVF-ET program. Methods: Frozen-thawed and survived human blastocysts were treated by immunosurgery, and recovered ICM cells were cultured onto STO feeder cell layer and ICM colony was subcultured by mechanical dissociation into clumps. To identify ES cell, alkaline phosphatase staining and expression of Oct4 in replated ICM colonies were examined. Also, to examine the possibility of ES cell differentiation, retinoic acid (RA), basic fibroblast growth factor (b-FGF), nerve growth factor (NGF) were added in culture medium. In addition, to classify the specific cell type, differentiated cells were stained by indirect immunocytochemistry. Results: One ICM colony recovered from frozen-thawed six blastocysts was subcultured, continuously replated during 40 passage culture duration without differentiation. Subcultured colonies were strong positively stained by alkaline phophatase. When the expression of Oct4 in cultured ES colony was examined, Oct4b type is more clearly indicated than Oct4a one although there was not detected in embryoid body or differentiated cells. In differentiated cardiomyocytes from ES colony, cells were beaten regularly (60 times/min). In differentiated neural cells from ES colony, neurofilament (NF) 200 kDa protein, microtubule associated protein (MAP) 2 and ${\beta}$-tubulin of specific marker in neurons, glial fibrillary acidic protein (GFAP) of specific marker in astrocytes and galactocelebrocide (GalC) of specific marker in oligodendrocytes were confirmed by indirect immunocytochemistry. Also, muscle cells were detected by indirect immunocytochemistry. In addition, ES colonies can be successfully cryopreserved. Conclusion: This study suggested that establishment of human ES cells can be successfully derived from frozen-thawed blastocysts that were destined to be discarded, and obtained specific cell types (cardiomyocytes, neurons and muscle cells) through the in vitro differentiation procedures of ES cells.

• Research in Plant Disease
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• v.19 no.3
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• pp.201-207
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• 2013

This research was performed to establish basic technology for Chinese cabbage clubroot chemical control by investigating the soil contamination of Plasmodiophora brassicae in major producing regions of fall Chinese cabbage. PCR primers were developed to detect P. brassicae, a causal agent of Chinese cabbage club-root that generally occurs in Cruciferae family. A primer set, PbbtgF761 and PbbtgR961, specifically amplified a 245 bp fragment from P. brassicae only. At places well known for fall Chinese cabbage, 10 out of 33 in Haenam-gun, 5 out of 13 in Yeongam-gun and Yeonggwang-gun, 1 out of 6 in Gochang-gun, 2 out of 12 in Hongseong-gun, and 5 out of 17 in Dangjin-si resulted positive for P. brassicae contamination. The results show that the soil contamination rate of P. brassicae was 30.3% in Haenam-gun, 38.5% in Yeongam-gun and Yeonggwang-gun, 16.7% in Gochang-gun, 16.7% in Hongseong-gun, and 29.4% in Dangjin-si. The six places where Chinese cabbage clubroot was visible by naked eye were 100% confirmed by the PCR test of the P. brassicae contaminated soil. Thus, simple PCR test may be utilized as an index to decide on chemical control of P. brassicae.

• Journal of Life Science
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• v.16 no.5
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• pp.729-733
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• 2006

Cryptosporidium is a waterborne pathogenic parasite which causes diarrhea. Immunomagnetic separation-immunofluorescent assay (IMS-IFA) has been a widely adopted for Cryptosporidium detection as standard method. However, this method does not provide information about viability or infectivity of Cryptosporidium. Therefore, many researchers have studied viability or infectivity analyses of Cryptosporidium with various methods such as vital staining, in vitro excystation, RT-PCR, cell culture, and mouse infection assay. In this study, two direct RT-PCR methods, cell culture RT-PCR and cell culture IFA were compared for sensitivity and other characteristics. The results showed that direct RT-PCR method with HSP70 genes had the highest sensitivity with detection up to 1 viable cell of Cryptosporidium. The infectious Cryptosporidium were detected up to 10 to 25 cells by cell culture methods in combination with RT-PCR and IFA. The infectious Cryptosporidium were apt to be quantified by cell culture IFA.

• Korean Journal of Microbiology
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• v.48 no.4
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• pp.284-292
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• 2012

To date, chemical control remains the most common way to reduce beet armyworm (Spodoptera exigua) populations. However, this insect has become more tolerant or resistant to many chemical insecticides and the insect larvae usually hide inside hollow, tube-like leaves of host plant so they were difficult to kill by spraying insecticides. The use of viral and bacterial insecticide to solve these problems has not been successful because of their novel feeding habit. To overcome these problems, in this study, the biological characteristics and virulence of an entomopathogenic fungus isolated from the cadaver of larvae beet armyworm were investigated. Isolated entomopathogenic fungus was identified as Nomeraea rileyi (Farlow) Samson by morphological examinations and genetic identification using sequences of the ITS, ${\beta}$-tubulin gene and EF1-${\alpha}$ regions. This fungus was named as N. rileyi SDSe. Virulence tests against 3rd larvae of beet armyworm were conducted with various conidial suspensions from $1{\times}10^4$ to $10^8$ conidia/ml of N. rileyi SDSe in laboratory conditions. Mortality rate of beet armyworm showed from 20 to 54% and the virulence increased with increasing conidial concentrations. Although N. rileyi SDSe showed low mortality rate against beet armyworm, it is expected that N. rileyi SDSe will be used effectively in the integrated pest management programs against the beet armyworm.