• Proceedings of the Plant Resources Society of Korea Conference
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• 2018.04a
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• pp.45-45
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• 2018

We had already reported the successful germination for green pods of purple lady's slipper orchid (Cypripedium macranthos Sw.). The green pod methods is to take immature seeds in green capsules, sterilize the capsule, and take out the sterile seeds. This method, however, needs very critical time of harvest. The critical time of seed harvest changes depending upon the species, condition of the specimen, and climatic influence, and the right time lies between 5 and 12 weeks after fertilization. In this study, the mature seeds were collected after 120-130 days with hand-polination of lady's slipper orchids. Mature seeds are usually dormant and it has to be overcome, either with hormone or storing the seeds near freezing for two or three months to break dormancy. The seeds were first surface sterilized with 70% ethanol and then transferred 1% NaOCl for 10-15 minutes, followed by rinses 3 times with sterilized distilled water. The cypripedium seeds consists of an embryo within a seed coat known as a testa. The testa is water repellent and the seed has a large air space between the embryo and testa so the seed tends to float on water. We had resolved the problems with vacuum pump to soak water into the testa before sterilization. The seeds were placed on liquid or agar solidified germination media. Cultures were incubated at $24{\pm}1^{\circ}C$ in dark. The seeds were germinated in 6-8 weeks in liquid suspension culture (germination percentage over 18%); however, the seeds on agar solidified media took more than 5 months to germinate and the germination percentage less than 5%. The most effective media for liquid culture was 1/4 strength Murashige and Skoog (MS) medium with 50 ml/l coconut water ($4brix^{\circ}$) at pH 5.8.

• The Korean Journal of Mycology
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• v.34 no.2
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• pp.112-118
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• 2006

To select efficient entomopathogenic fungal strains of Lecanicillium for the biocontrol of aphid, Myzus persicae, conidial suspension ($1{\times}10\;conidia/ml$) was sprayed onto a detached Chinese cabbage leaf in a petri dish with a dampened filter paper that had 20 nymphs of aphid. Lecancillium strain 4078 and 6543 were the best strains for the biocontrol of aphid at high temperature of $30^{\circ}C$ and low relative humidity (RH) of 85%, respectively. The cumulative mortality of strain 4078 at $30^{\circ}C$ after 3 days was 100% and that of strain 6543 was 90% at 85% RH after 5 days. Strain 4078 also exhibited almost 100% germination ratio of conidia and high rate of mycelial growth at the broad temperature-range of $15{\sim}25^{\circ}C$. The strain 4078 and 6543 were all identified as Lecanicillium species based on the DNA sequences (accession no.: EF026004 and EF026005, respectively) of the ITS regions of the fungi. Excellent production of aerial conidia of strain 6543 was accomplished by using steamed polished rice as the solid culture medium.

• Korean Journal of Plant Tissue Culture
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• v.28 no.2
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• pp.103-108
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• 2001

The in vitro plantlets of cassava (Manihot esculenta Crantz cv. MColl 22) could be regenerated from nodal explant cultures in a liquid MS basal medium containing 0.01 mg/L zeatin for 2 weeks. The plantlets of 1.5∼2.5 cm in shoot length were transplanted to a glass bottle containing fine sand and acclimated under non-sterile conditions after excising their intact roots by: 1) prune leaving roots base of 1∼1.5 cm; 2) complete removal of roots; and 3) cutting off the rooting zone. The majority of in vitro-formed intact roots continued growth after transferred to soil, and all of the damaged roots stopped further growth. The plantlets with excised roots began to develop new roots within 7∼10 days after being transferred to a glass bottle, and a few of the pruned roots developed lateral roots from the remaining portion. Pruning and removal of in vitro roots resulted in a high survival rate (>87%), and did not significantly affect ex vitro root regeneration and acclimation, but the plantlets in which the rooting zone had been cut-off showed 73% survival rate. Pruning or removal of in vitro roots before transfer of plantlets is recommended for useful method of commercial micropropagation because of easier handling and high survival rate of plantlets.

• Asia-Pacific Journal of Business Venturing and Entrepreneurship
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• v.18 no.1
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• pp.225-236
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• 2023

Policy implications were derived by comparing/analyzing innovative SMEs and general SMEs that obtained innovation certification from 2015 to 2021 in terms of survival period. Work experience, scale (employment, capital and debt size, sales and operating profit) Korean standard industry classification (2 digit) was used to select general SMEs similar to innovative SMEs. Survival period was calculated by defining suspension, closure and overdue equivalent to default as events. As a result of the survival analysis, innovative SMEs showed a 9.8% reduction in the risk of delinquency compared to general SMEs, indicating that the survival period of innovative SMEs was significantly longer. In addition, it was found that the work experience and size (employment, capital) of SMEs had a positive effect on the survival period, but debt had a negative effect on the survival period. This means that the innovation certification system centered on innovation capabilities and future growth potential is a significant indicator in terms of survival period. As a result, it was concluded that the benefits and support policies provided by the innovation certification system need to be more systematic and sophisticated by reflecting the work experience and industry for the actual growth and survival of SMEs.

• Journal of Periodontal and Implant Science
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• v.26 no.4
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• pp.949-965
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• 1996

The main goal of periodontal therapy is to restore the lost periodontal tissue and establish the attachment appratus. Current acceptable therapeutic techniques are included : removal of diseased soft tissue, demineralization of exposed root surface, using the barrier membrane for preventing the downgrowth of gingival epithelial cell, insertion of graft materials as a scaffolding action, and biological mediators for promoting the cell activity. The latest concept one among them has been studied which based on the knowledge of cellular biology of destructed tissue. Platelet-derived growth factor(PDGF) is one of the polypeptide growth factor which have been reported as a biological mediator to regulate activities of wound healing progress including cell proliferation, migration, and metabolism. The purposes of this study is to evaluate the influences of the PDGF as biological mediator to periodontal ligament and bone marrow cell. Both right and left maxillary first molar were extracted from rat which had treated with 0.4% ${\beta}-Aminopropionitril$ for 5 days, and feeded until designed date to sacrifice under anesthesisa. Periodontal ligament were removed from the extracted socket of the rat, and cultured with Dulbecco's Modified Essential Medium(DMEM) contained with 10% Fetal Bovine Serum, 100U/ml penicillin, $100{\mu}g/ml$ streptomycin, $0.5{\mu}g/ml$ amphotericin-B. Bone marrow cell were culture from bone marrow suspension with which washed out from femur with same medium. The study was performed to evaluate the effect of PDGF to periodontal ligament and bone cell, cell proliferation rate, total protein synthesis, and alkaline phosphatase activity of rat periodontal ligament(PDL) cell and bone stromal(RBS) cell in vitro. The effects of growth factors on both cells were measured at 3, 5th day after cell culture with (control group) or without growth factors(experimental group). The results were as follows: 1. The tendency of cell proliferation under the influence of PDGF showed more rapid proliferation pattern than control at 3 and 5 days after inoculation. 2. The activity of Alkaline phosphatase revealed 14, 80% increased respectively at 3, 5 days culture than control group. Measurements of ALPase levels indicated that PDL cells had significantly higher activity when compared with that of co-culture groups and GF only(P<0.05). And, ALPase activity in 10 days was higher than that of 7 days(P<0.05). 3. The tendency of formation of the mineralized nodule were observed dose-depend pattern of PDL cells. There was statistically significant difference among group l(PDL 100%), 2(PDL 70%:GF 30%), and 3(PDL 50%:GF 50%)(P<0.01). But, there was no difference among group 3, 4(PDL 30%:GF 70%), and 5(GF 100%). 4. Also, the number of nodule was greater in co-culture of PDL 70% and GF 30% than in culture of PDL 70%(P<0.05). From the above results, it is assumed that the PDGF on PDL cells and RMB cell culture. GF stimulates the cell growth, which is not that of PDL cells but GF. And, the activity of ALPase depends on the ratio of PDL cells, and ALPase may relate to the initial phase of nodule formation. Also, it is thought that the calcified nodule formation principally depends on PDL cells, is inhibited by GF, and affected by cell density. In conclusion, platelet-derived growth factor can promote rapid osteogenesis during early stage of periodontal tissue regeneration.

• Korean Journal of Poultry Science
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• v.40 no.3
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• pp.197-205
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• 2013

This study established a method for preserving chicken primordial germ cells (PGCs) that enables long-term storage in liquid nitrogen ($LN_2$) for preservation of the species. The purpose of this study was to compare the effects of Ethylene Glycol (EG) and Propylene Glycol (PG) on viability of cryopreserved PGCs with vitrification in Korean Native Chicken (Ogye), and to fine should be find or to the optimal protocol for PGCs freezing. One of the important components of cryopreservation process is cryopreservation medium that plays a vital role in preventing cellular injury during freeze-thawing. Cryoprotective agents have been known to improve cell viability after freeze-thawing. PGCs obtained from the germinal gonade of 5.5~6 day (stage 28) chick embryos, using the MACS method were suspended in a freezing medium containing a freezing and protecting agents. Gonads were harvested from stage 28 chick embryos and pooled in groups of 10E embryos, contributing gonads to the cell suspension. The gonadal cells, including PGCs, were then frozen in 1 of the following cryoprotectant treatments: 2.5% EG, 5% EG, 10% EG, 2.5% PG, 5% PG, 10% PG, and 0% cryoprotectant as a control. Effects of exposure to vitrification solution and vitrification, with different concentrations of the cryoprotectant solution, were examined. After freezing and thawing, survival rates of the frozen-thawed PGCs from the 0, 2.5, 5, 10 and 15% EG plus FBS treatment were 44.24%, 64.51%, 85.63%, 80.51% and 73.52% (p<0.05), respectively. The viability of PGCs after freeze-thawing was significantly higher for 10% EG plus FBS treatment than for 10% PG + FBS treatment (p<0.05)(85.63% vs 66.81%). Therefore, these systems may contribute in the improvement of cryopreservation for a scarce species in birds preservation. This study established a method for preserving chicken PGC that enables systematic storage and labeling of cryopreserved PGCs in liquid N at a germplasm repository and ease of entry into a database. In the future, the importance for this new technology is that poultry lines can be conserved while work is being conducted on improving the production of germline chimeras.

• FLOWER RESEARCH JOURNAL
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• v.18 no.1
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• pp.1-8
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• 2010

Sensitivities of PLBs of four Phalaenopsis cultivars, P. 'Taisuco Windian', P. 'Nancy Amour', P. 'Pink Twilight' and P. 'Taipei Gold' to kanamycin, spectinomycin and hygromycin at different concentrations (0, 25, 50, 100, 200, and $400mg{\cdot}L^{-1}$) were examined. Hygromycin was favorable for selecting the transformants in the genetic transformation of Phalaenopsis as PLBs of four cultivars were all dead at even $25mg{\cdot}L^{-1}$ hygromycin. Responses of PLBs of P. 'Maki Watanabe' and P. 'Brother Lawrence' to DL-phosphinothricin (PPT) were determined at different concentrations (0, 0.1. 0.25, 0.5, 1.0, 2.0, 2.5, and $5.0mg{\cdot}L^{-1}$) and $0.5mg{\cdot}L^{-1}$ PPT was thought to be suitable for selecting the transformants of Phalaenopsis. The optimum conditions for Agrobacterium cocultivation with Phalaenopsis PLBs were examined using a two-step cocultivation method in Dtps. 'City Girl' and A. tumefaciens LBA4404. In the first infection period in a 1 : 10 suspension of Agrobacterium to a VW medium, 1 hr infection showed the highest PLB survival ratio. And then, PLBs were cocultivated with a bacterial strain and a 3-day cocultivation period was better for Phalaenopsis PLBs than a prolonged period. Agrobacterium tumefaciens strains LBA4404 (pTOK233) and EHA105 (pGA643) were used to compare their efficiency on the genetic transformation of Phalaenopsis PLBs. The PLBs infected with EHA105 survived more than those infected with LBA4404 after two days in a dark condition and two weeks in light condition on a selective medium. About 1,000 PLBs for each of P. 'Maki Watanabe' and P. 'Brother Lawrence', and each bacterial strain of AGL1 (pCAMBIA3301) and LBA4404 (pTOK233) were used for the regeneration of transgenic plants. The bacterial strain AGL1 had a higher genetic transformation efficiency than LBA4404, with no significant difference between cultivars. In this study, 11 hygromycin-resistant plantlets and 32 PPT-resistant plantlets were produced, but these putative transgenic plantlets need further examinations.

• Korean Journal of Soil Science and Fertilizer
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• v.40 no.6
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• pp.447-453
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• 2007

With a broad objective for the development of microbial based fertilizers, a total of 373 strains were isolated from rhizoplane and rhizosphere of pepper, tomato, lettuce, pasture, and grass. The efficacy of the isolates to augument overall plant growth was evaluated. After screening for their plant growth promotion and antagonistic properties in vitro efficient strains were further selected. The most efficient strains was characterized by 16S rRNA gene sequences and biochemical techniques and was designated as Bacillus subtilis S37-2. The strains facilitated plant growth and inhibited the plant phathogenic fungi such as Fusarium oxysporum (KACC 40037, Rhizoctonia solani (KACC 40140), and Sclerotinia sclerotiorum (KACC 40457). Pot based bioassay using lettuce as test plant was conducted by inoculating suspension ($10^5$ to $10^8cells\;mL^{-1}$) of B. subtilis S37-2 to the rhizosphere of lettuce cultivated in soil pots. Compared with non-inoculated pots, marked increase in leaf (42.3%) and root mass (48.7%) was observed in the inoculation group where the 50ml of cell mixture ($8.7{\times}10^8cells\;ml^{-1}$) was applied to the rhizosphere of letuce either once or twice. Antagonistic effects of B. subtilis S37-2 strain on S. sclerotiorum (KACC 40457) were tested. All the tested lettuce plants perished after 9 days in treatment containing only S. sclerotiorum, but only 17% of lettuce was perished in the inoculation plot. B. subtilis grew well in the TSB culture medium. The isolates grew better in yeast extracts than peptone and tryptone as nitrogen source. The growth rate was 2~4 times greater at $37^{\circ}C$ as compared with $30^{\circ}C$ incubation temperature. B. subitlis S37-2 produced $0.1{\mu}g\;ml^{-1}$ of IAA (indole 3-acetic acid) in the TSB medium containing L-tryptophan($20mg\;L^{-1}$) in 24 hours.

• Korean Journal of Animal Reproduction
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• v.21 no.2
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• pp.167-176
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• 1997

This study was carried out to evaluate the potential for preselection of transgenic embryos prior to transfer into recipient animals. In these experiments, I used a 3.2 kb transgene which contained the neomycin resistance gene (neo) and lac Z gene driven by the $\beta$ actin promoter (iacZ Ineo). Oocytes were aspirated from abattoir ovaries, matured in TCM-199 supplemented with 10% fetal bovine serum (FBS), 5 ${\mu}\textrm{g}$/ml LH, 0.5 ${\mu}\textrm{g}$/ml FSH, 100 unit/ml penicillin, and 100 ${\mu}\textrm{g}$/ml streptomycin for 22 to 24 hrs then inseminated with a sperm suspension of 1 X 10$^6$ sperm/ml containing 5 ${\mu}\textrm{g}$/ml of heparin. At 18-20 hrs after insemination, cumulus cells were removed by vortexing and pronuclei of centrifuged zygotes microinjected with the lacZ/neo construct (3 ng/$\mu$l). All cultures were carried out in CR1aa with transfected BRL monolayers containing 3 mg/ml BSA, 20 $\mu$/ml NEM amino acids and 40 $\mu$I/ml BME amino acids. To identify the appropriate concentration of G418 for selection, non-microinjected zygotes were cultured in the presence of 0, 50, 100 and 200 $\mu$g/ml of G418. After 8 days of culture in these treatments, 44/145 (30.3%), 13/150 (8. 7%), 1/151 (0.7%) and 0/134 (0.0%) devel-oped to the blastocyst stage in 0, 50, 100 and 200 $\mu$g/ml of G418, respectively. A total of 1,127 zygotes were microinjected and placed into culture (without G418) and subsequently 710 (63.0%) cleaved. At 48 hrs post-injection, embryos ($\geq$2-cell) were randomly assigned to control (medium alone) or G418 (100 ${\mu}\textrm{g}$/ml) treatments. A control culture of 740 non-microinjected embryos from the same replicates of embryos were also placed into control medium. After 8 days in culture, 54/343 (15.7%) and 22/367 (6.0 %) of the microinjected embryos developed to the blastocyst stage in control and G418 media, respectively. A total of 151/740 (27.2%) of the non-microinjected embryos placed in the control medium developed to the blastocyst stage. The blastocysts in the control treatment had a mean of 70.7 ${\pm}$ 4.7 cells of which 23.1 ${\pm}$ 2.6 (32.7%) stained for $\beta$-Gal activity. B1astocysts in the G418 treatment had a mean of 48.8${\pm}$7.5 cells of which 40.3 ${\pm}$ 4.1 (82.6%) stained for $\beta$-Gal ac tivity (P<0.05). In the control treatment 26 of 30 (87.0%) blastocysts had some cells with $\beta$-Gal activity while all of the blastocysts in the G418 treatment had some cell with $\beta$-Gal ac tivity (P<0.05). However, ICM colonies in either control or G418 treatments were not expressed any epiblast cell except of trophetoderm celIs. The $\beta$-actin promoter/lacZ gene may not be e expression or silence expression in epiblast cells These results clearly show an enrichment of blastocysts expressing the transgene in the majority of their cells after culture in the presence of G418. The exogeneous gene was not express a and silence in ICM colonies especiallly epiblast cells except of trophectederm cells. Even though the higher rate cell number expressed of exogeneous gene in the G418 treatments, a total cell number was decrease significantly (P<0.05) of which might be also drop of the establishment of ES like-cell colonies and production of transgenic animals. However, futher studies need to determine the viability of these selected embryos and the avability of production of transgenic offspring.

• Horticultural Science & Technology
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• v.33 no.3
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• pp.409-419
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• 2015

This study was conducted to establish an efficient screening method for watermelon plants resistant to Fusarium wilt (FW), which is caused by Fusarium oxysporum f. sp. niveum (Fon). An HA isolate was prepared from a wilted watermelon plant in Haman-gun and identified as F. oxysporum f. sp. niveum based on morphological characteristics, molecular analyses of ITS (internal transcribed spacer) and TEF (translation elongation factor $1{\alpha}$) sequences, and host specificity on cucurbits including watermelon, melon, oriental melon, and cucumber. The assay for disease response of watermelon differentials indicated that the HA isolate was race 0. Among seven liquid media tested, the highest amount of Fon spores was produced from V8-juice broth, which was selected as a medium for mass production of Fon. The disease assay for 21 watermelon and 11 watermelon-rootstock cultivars demonstrated that 20 watermelon cultivars except for 'Soknoranggul' were susceptible; 'Soknoranggul' was moderately resistant. All the tested rootstock cultivars were highly resistant to the HA isolate. The evaluation of disease development depending on various conditions suggested that an efficient screening method for FW resistance in watermelon plants is to dip the roots of 10-day-old seedlings in spore suspension of $1.0{\times}10^5-1.0{\times}10^6conidia{\cdot}mL^{-1}$ for 30 min., to transplant the seedlings to plastic pots with a fertilized soil, and then to cultivate the plants at $25^{\circ}C$ for 3 weeks.