Insect pollinators of the endanger orchid Cypripedium japonicum were surveyed and identified during two years, as a part of a conservation project of the orchid at Jukyeup-san and Hwaak-san (Mt.), South Korea. In total 40 individuals of 16 species in 4 families were identified. The dominant family was Halictidae, and Lasioglossum exiliceps Vachal visited the most frequently C. japonicum during the surveys. The average visiting frequency was 2.5 individuals per hour and the highest 4.3, from 12:00 – 13:00 in a day. After 15:00 insects did not visit the flowers at all. However, all of the visiting insects were found to not carry a pollinium or pollens of the orchard on their bodies; pollen carryover by any of the visiting insects did not occur at all. The orchid seems to require certain pollinators in particular body thickness due to its unique pollination mechanism. The orchid has two exit route openings, around 1 cm in diametre, where the entrapped insects can exit and an anther is situated just in front of each opening. It was inferred that a pollen carrier should be around 1 cm in body thickness. Therefore, the candidate species as the proper pollen carriers can be Tetralonia nipponensis Perez, Xylocopa appendiculata circumvolans Smith and Bombus consobrinus Dahlbom among the surveyed visitors.

Germplasm collection of lily in the mountainousarea is laborious and time consuming accompanied withhigh costs and high risk due to inappropriate environmentalconditions. Cryopreservation being an ideal method for thelong-term preservation can be employed in conservation ofvaluable lily germplasm. Previously, we developed cryo-preservation protocol for Lilium germplasm using ‘dropletvitrification’. In this paper, we have chosen shoot tips as amaterial for cryopreservation because of their genetic safetyupon regrowth in tissue culture. Using this protocol, wehave preserved approximately 160 accessions of lily germ-plasm in 2010~2012. The regeneration rates are rangedfrom 54.3% to 58.5% while the survival rates were from58.3% to 66.4%. Among Lilium germplasm cryopreserved,there are some Korean, Chinese, and Taiwanese seedstocks which have good qualities for inter-species hybrid.Moreover, we also conserved Korean wild endangered seedstock, especially Lilium hansonii. The morphological studyof Lilium germplasm regenerated from cryopreserved mate-rial confirmed the stability of clonal material following cryo-preservation. We anticipate this cryo-collection will beavailable and useful to curators or breeders of Lilium andthis cryobank will also facilitate the conservation and inter-national exchange of Lilium germplasm.

Polyhedrin is the major component of the nuclear viral occlusions produced during replication of the baculovirus Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV). To enhance the production efficiency of foreign protein in baculovirus expression system, the effects of various polyhedrin fragments were investigated by fusion expressing them with the enhanced green fluorescent protein (EGFP). Recombinant viruses were generated to express EGFP fused with polyhedrin fragments based on the previously reported minimal region for self-assembly and the KRKK nuclear localization signal (NLS). The marked increase of EGFP by these fusion expressions was confirmed through protein and fluorescence intensity analyses. Among the fusion-expressed protein in nucleus and cytoplasm, the most hyper-expression was observed in the fusion of amino acids 19 to 110 and 32 to 59 of polyhedrin. Also these fragments, some degradation of only the fused polyhedrin was observed in the fusion of amino acids 19 to 85 and 32 to 85. The production of E2 protein, which is a major antigen of classical swine fever virus, was dramatically increased by fusion expression with polyhedrin amino acids 19 to 110, and its preliminary immunogenicity was verified using experimental guinea pigs. The production of luciferase was approximately two folds increased by fusion expression with polyhedrin amino acids 32 to 59, and its activity was measured using Luminometer. This study suggests a new option for higher expression of useful foreign recombinant protein using the partial polyhedrin fusion expression in baculovirus.

Flexible transparent conducting films (TCFs) were fabricated by dip-coating single-wall carbon nanotubes (SWCNTs) onto a flexible polyethylene terephthalate (PET) film. The amount of coated SWCNTs was controlled simply by dipping number. Because the performance of SWCNT-based TCFs is influenced by both electrical conductance and optical transmittance, we evaluated the film performance by introducing a film property factor using both the number of interconnected SWCNT bundles at intersection points, and the coverage of SWCNTs on the PET substrate, in field emission scanning electron microscopic images. The microscopic film property factor was in an excellent agreement with the macroscopic one determined from electrical conductance and optical transmittance measurements, especially for a small number of dippings. Therefore, the most crucial factor governing the performance of the SWCNT-based TCFs is a SWCNT-network structure with a large number of intersection points for a minimum amount of deposited SWCNTs.

The purpose of this study was to test the effect of Gastrocnemius and Low Back-muscle isotonic exercise on static•dynamic standing balance during the period of 4 weeks. This study was two groups pretest-posttest design. Nineteen subjects who were over 22 years old were randomly assigned to either the experimental group that received the Gastrocnemius muscle exercise(n=9) or the low back muscle exercise(n=10) : The former group performed isotonic exercise(plantar flexion), the latter group performed isotonic exercise(trunk extension) a total of 18 times for three times per week for four weeks. Two groups also performed static and dynamic balance before the exercise and 4weeks after the exercise. The data were analyzed by using the paired t-test and independent t-test. The results were as follows: As compared with change of dynamic balance performance capacity at two groups, a significant difference was shown in the test(p<.05), but not in static balance(p>.05). Also, a significant difference of balance between groups was not shown in the test. In this study indicated that gastrocnemius and low back muscle isotonic exercise will have positive impact on standing balance.

The fact that flip-flops, one of many different types of unstable shoes, are light and relatively easy to put on, accounts for their popularity among people. But because flip-flops rely heavily on the support of a single thong between your first and second toes, they impose a huge amount of pressure onto lower leg. Thus in the following experiment we tried to examine the different effects of flip-flops and running shoes in terms of their effect on muscle activity and fatigue of tibialis anterior and gastrocnemius during walking. In order to measure an electromyogram we used Free EMG system. 10 men and 10 women in running shoes ran on treadmills for 15 minutes at 4.8km/h, 2 days later the same experiment was carried out, but this time, in flip-flops. p value turned out to be greater than .05 and thus there was no considerable difference between the effects of flip-flops and running shoes on muscle activity and fatigue during walking. Therefore we conclude that despite the fact that flip-flops are considered unstable, their effects on muscle activity and fatigue of tibialis anterior and gastrocnemius are negligible.

Apolipophorin III (apoLp-III) is a well-known hemolymph protein having a functional role in lipid transport and immune response of insects. We cloned full-length cDNA encoding putative apoLp-III from larvae of the coleopteran beetle, Tenebrio molitor (TmapoLp-III), by identification of clones corresponding to the partial sequence of TmapoLp-III, subsequently followed with full length sequencing by a clone-by-clone primer walking method. The complete cDNA consists of 890 nucleotides, including an ORF encoding 196 amino acid residues. Excluding a putative signal peptide of the first 20 amino acid residues, the 176-residue mature apoLp-III has a calculated molecular mass of 19,146 Da. Genomic sequence analysis with respect to its cDNA showed that TmapoLp-III was organized into four exons interrupted by three introns. Several immune-related transcription factor binding sites were discovered in the putative 5’-flanking region. BLAST and phylogenetic analysis reveals that TmapoLp-III has high sequence identity (88%) with Tribolium castaneum apoLp-III but shares little sequence homologies (<26%) with other apoLp-IIIs. Homology modeling of Tm apoLp-III shows a bundle of five amphipathic helices, including a short helix 3’. The ‘helix-short helix-helix’ motif was predicted to be implicated in lipid binding interactions, through reversible conformational changes and accommodating the hydrophobic residues to the exterior for stability. Highest level of TmapoLp-III mRNA was detected at late pupal stages, albeit it is expressed in the larval and adult stages at lower levels. The tissue specific expression of the transcripts showed significantly higher numbers in larval fat body and adult integument. In addition, TmapoLp-III mRNA was found to be highly up-regulated in late stages of L. monocytogenes or E. coli challenge. These results indicate that TmapoLp-III may play an important role in innate immune responses against bacterial pathogens in T. molitor.

Mealworms, Tenebrio molitor (L.) is used as an important animal feed additive for growth promotion and health management, but potentially exposes to fungal infection. In this work, virulence of two species of entomopathogenic fungi against the insect, and the relationship between abiotic features and virulence were investigated. Secondly our consideration was also given to the effect of chemical fungicides on conidial germination for risk control. Between Beauveria bassiana (Bb) and Metarhizium roberstii (Mr) (previously M. anisopliae), Bb isolates had much higher virulence (~100% mortality in 3~4 days after the treatment), rather than Mr isolates in laboratory assays. Next, fungus-treated mealworms were kept at wheat bran at 20, 25, 30 and 35℃ with 3, 6, 9 times of water spray to the feeds for set-up of different humidity conditions. Inoculation of fungi to mealworms was conducted by fungal spray and feeding methods, which resulted in higher virulence in feeding method. In the feeding method, all temperature treatments except 35℃ showed high virulence against mealworms, but any significant relationship between virulence and humidity was not observed. In the chemical fungicide screening, fluazinam (CAS No. 79622-59-6) and mancozeb (8018-01-7) significantly inhibited the germination of Bb and Mr conidia. This work suggest that contamination of wheat bran with fungal pathogens, particularly B. bassiana may induce mycosis of mealworms, but introduction of effective fungicides possibly reduce fungal infection.

To enhance the production efficiency of foreign protein in baculovirus expression system, the effects of polyhedrin fragments were investigated by fusion expression them with the enhanced green fluorescence protein (EGFP). Recombinant viruses were generated to express EGFP fused with polyhedrin fragments based on the minimal region for self-assembly and the KRKK nuclear localization signal (NLS). The increase of EGFP production by fusion expressions was confirmed through protein and fluorescence intensity analyses. The importance of nuclear localization for enhanced production of EGFP was shown by the mutation of the NLS within the fused polyhedrin fragment. Among the fusion expressed protein in cytoplasm, the most hyper-expression was observed in the fusion of amino acids 32 to 59 of polyhedrin. Polyhedrin fragment fusion expression with classical swine fever virus E2 protein also resulted hyper-enhanced expression of E2 protein. However, the fusion expression of porcine circovirus ORF2 with polyhedrin fragment did not show significant enhance of ORF2 production. These results suggested that the enhancement of foreign protein production when fused with polyhedrin is caused by the enhanced stability of expressed protein.

Shitao(:tJ11) suggested that the one stroke of painting is the beginning of the Universe. he claimed that the one is the foundation of all the images. It is based on the DaoJ! that is erupted myriad creatures. His theory of one stroke is showing the root of the world and mind of artists. His theory is a mind theory of mind as well. As a Korean painter, I would like to explain my painting by the Shitao's theory of one stroke. It is painted by natural color from pine tree to draw the pine tree. It is claimed that one stroke is displayed by the way of seamless from the material to art.

This study aims to examine the effects of taping of the ankle joint on the static and dynamic balance and gait ability of stroke patients. Twenty-six stroke patients receiving physical therapy at a hospital located in Gyeonggi-do were divided equally into a group that had taping in physical therapy and an ordinary physical therapy group. They exercised for 30 minutes each, 3 times per week for 8 weeks from June to August 2011. Romberg’s eye open and eye closed tests, limits of stability(LOS), forward and back test, timed up and go test(TUG) and 10-meter gait velocity test were performed to evaluate static balance, dynamic balance, and gait ability, respectively, prior to and 8 weeks after the intervention. Differences within each group in relation to the lapse of time were compared by a paired t-test. Differences between the two groups were compared by an independent t-test. Regarding comparison of differences within each group, all tests resulted in significant changes in both groups after the intervention (p<.05). Comparison of differences between the two groups showed that taping in the physical therapy group had significantly better test results than the ordinary physical therapy group in all measured items(p<.05). The after effects of ankle taping on stroke patients are more efficient and effective than ordinary physical therapy alone in improving balance and gait ability.

Interferon induced transmembrane protein-1 (Ifitm-1) has been reported to have an important role in primordial germ cell formation, and it has expressed in female reproductive organ. In the present study, Ifitm-1 gene expression was identified in testes and all part of epididymis using western immunoblot and immunohistochemistry. Interestingly, Ifitm-1 expression was observed on the head of spermatozoa. To investigate the role of Ifitm-1 gene expression in behavior of spermatozoa after acrosome reaction, fresh sperm was incubated with calcium ionophore to induce acrosome reaction, whereas the expression of Ifitm-1 was not altered after the acrosome reaction. Then to identify the effect of Ifitm-1 in sperm motility and other seminal parameters, different concentration of Ifitm-1 antibody was incubated with spermatozoa, and seminal parameters were assessed using computer-assisted semen analysis (CASA). Interestingly, motility, progressive, and VAP were increased in the sperm with Ifitm-1 antibody treated compared to rabbit serum, however other parameters such as straightness were not changed. In order to identify the functional significance of Ifitm-1 in fertilization, capacitated spermatozoa were pre-incubated with anti- Ifitm-1 antibody and subsequently examined the ability to adhere to mouse oocytes. However, any defection or alteration in sperm-egg fusion was not found, Ifitm-1 antibody treated or non-treated spermatozoa showed a normal penetration. Although the precise role of Ifitm-1 in sperm motility and following fertilization need to be elucidated, this study suggests that the activation of Ifitm-1 on the sperm may enhance the motility of spermatozoa in mice.

The T-cell receptor (TCR) engages with an antigen and initiates a signaling cascade that leads to the activation of transcription factors. Roquin, a protein encoded by the RC- 3H1 gene and characterized as an immune regulator, was recently identified as a novel RING-type ubiquitin ligase family member, but the mechanisms by which Roquin regulates T-cell responses are unclear. We used the EL-4 murine lymphoma cell line to elucidate the role of Roquin in vitro. Roquin-overexpressing EL-4 cells became hyper-responsive after anti-CD3/CD28 stimulation in vitro and were a major source of the cytokines IL-2 and TNF-α. Upon activation, these cells showed particularly enhanced production of IL-2 and TNF-α. To clarify the important role played by Roquin in T-cell responses ex vivo, we generated T-cell-specific Roquin transgenic (Tg) mice. Roquin-Tg CD4+ T-cells showed enhanced production of IL-2 and TNF-α in response to TCR stimulation with anti-CD28 co-stimulation. Further studies are necessary to investigate the role of Roquin in the regulation of primary T-cell activation, survival, and differentiation.

Pluripotent stem cells can be derived from both pre- and post-implantation embryos. Embryonic stem cells (ES cells), derived from inner cell mass (ICM) of blastocyst are naïve pluripotent and epiblast stem cells (EpiSCs) derived from post-implantation epiblast are primed pluripotent. The phenotypes and gene expression patterns of the two pluripotent stem cells are different each other and EpiSCs thought to be in a more advanced pluripotent (primed pluripotent state) than mouse ES cells (naïve pluripotent state). Therefore, we questioned whether EpiSCs are less potential to be differentiated into specialized cell types in vitro. EpiSCs were isolated from 5.5～6.5 day post coitum mouse embryos of the post-implantation epiblast. The EpiSCs could differentiate into all tree germ layers in vivo, and expressed pluripotency markers (Oct4, Nanog). Interestingly, EpiSCs also were able to efficiently differentiate into neural stem cells (NSCs). The NSCs differentiated from EpiSCs (EpiSC-NSCs) expressed NSC markers (Nestin, Sox2, and Musasi), self-renewed over passage 20, and could differentiate into two neural subtypes, neurons, astrocytes and oligodendrocytes. Next, we compared global gene expression patterns of EpiSC-NSCs with that of NSCs differentiated from ES cells and brain tissue. Gene expression pattern of brain tissue derived NSCs were closer to ES cell-derived NSCs than EpiSC-NSCs, indicating that the pluripotent stem cell-derived somatic cells could have different characteristics depending on the origin of pluripotent stem cell types. * This work was supported by the Next Generation Bio-Green 21 Program funded by the Rural Development Administration (Grant PJ 008009).

True hermaphrodites are animals of equivocal sex in which both male and female gonads develop simultaneously. The frequency of true hermaphroditism is higher in pigs than in other domestic animals. Two Korean pigs were diagnosed with true hermaphroditism showing ovotestes, epididymes, penes, and uteri. Histomorphologically, the testicular tissues consisted of Sertoli cells that were devoid of spermatogenic germ cells and showed proliferation of interstitial cells. However, the uteri were of normal architecture and had well-developed uterine endometrial glands. The samples were 38, XX female karyotype without the sex-determining region Y (SRY) gene. The findings of this study could contribute to the understanding of true hermaphroditism in the Korean pig industry. * This work was supported by a grant (Code# PJ008148) from BioGreen21 Program, Rural Development Administration, Republic of Korea.

The aim of this study was to examine the effect of acteoside (the cyclin-dependent kinase inhibitor) on the SCNT efficiency with adult fibroblasts in dog. Canine adult fibroblasts were obtained from muscle and cell cycle of fibroblasts was synchronized by culturing to confluency, serum starvation and treating with 30 μM acteoside for 48 h. Cell cycle stages, cell cytotoxicity (apoptosis) and, prduction of reactive oxygen species (ROS) were analyzed using flow cytometry. The canine cells, prepared by confluent-cell culture or treating with 30 μM acteoside for 48 h, were injected into enucleated in vivo matured oocytes, the couplets were electrical fused and activated by calcium ionomycin. SCNT embryos using acteoside-treated fibroblasts were surgically transferred into oviducts of estrus cycle synchronized recipient dogs. In cell cycle synchronization (G0/G1), there was no significant difference between serum starvations (83.9%) and acteoside treated groups (81.3%) that were higher than confluent group (78.5%). In production of apoptosis, confluent and acteoside treated groups (4.3 and 4.5%, respectively) were generated less than serum starvation group (21.8%). In case of ROS, serum starvation group was induced a significantly higher than other groups. After synchronization of the donor cell cycle, either confluent or acteoside treated, cells were placed with enucleated in vivo-matured dog oocytes, fused by electric stimulation, activated, and transferred into naturally estrus-synchronized surrogates. Fusion and cleavage rate of acteoside treated group were 64.1 and 41.5%, which were higher than those of confluent group (53.9 and 20.6%, respectively). The reconstructed embryo development rates to 4-cell and 8-cell in acteoside treated group were 29.5 and 14.8%, respectively, while confluent group showed 11.1 and 3.2%, respectively. Total 54 SCNT embryos using acteoside-treated fibroblasts were transferred into oviducts of 2 recipient dogs and one recipient finally delivered one puppy, whereas din`t detected pregnancy on transfer of cloned embryos reconstructed with confluent cells in 6 surrogate dogs. In conclusion, the results of the current study demonstrated that canine fibroblasts could be successfully arrested at the G0/G1 stage with reduced the formation of ROS and apoptosis after acteoside treatment. This results may contribute to improve the effi-ciency of canine SCNT. * This research was supported by iPET (Grants 110056-3), Ministry for Food, Agriculture, Forestry and Fisheries, Republic of Korea.

Acteoside acts as an anti-oxidative activity and anti-apoptosis in the cells. But, it has been not studied on maturation and development of porcine oocytes. The aims of the present study were to examine the effects of acteoside on the morphological progress of meiosis, developmental competence, and ROS in porcine oocytes. Oocytes were matured in tissue culture medium-199, supplemented with acteoside at various concentrations: 0 (control), 10, 30 and 50 μM. The oocytes maturation rates of groups supplemented with acteoside were no significantly different (81.13, 85.96, 82.95 and 83.68%, respectively). Level of ROS was significantly decreased in acteoside treated group. Furthermore, the parthenogenetic blastocyst rate was significantly improved in 10 μM acteoside treated group compared with control group (44.83 vs. 27.75%). And we investigated effect of acteoside on the oocytes condition represented by cytoplasmic maturation by homogeneous distribution and formation of cytoplasmic organelles and regulation of apoptosis-related genes. In the results. during IVM, 10 μM acteoside treated oocytes showed that the mitochondria and lipid droplet were smaller and homogeneous distribution in cytoplasm compare with control oocytes. And reverse transcription polymerase chain reaction (RTPCR) of parthenogenetic blstocysts revealed that acteoside increased the anti-apoptotic genes (Mcl-1, Bcl-2 and Bcl-xL), whereas reduced the expression of pro-apoptotic genes (Bax and Bak). In conclusion, based on the results, the effect of acteoside on IVM was not attractive. However, in acteoside treated group, cytoplasmic maturation seemed to be improved with morphologically uniform distribution of cytoplasmic organelles. Furthermore, embryonic development in acteoside treated group was significantly highly increased than that of non-treated group. Our results represents that addition of acteoside to the IVM medium has a beneficial effect in physiology of porcine oocytes, providing a improved method for porcine oocytes in vitro. * This work was supported by a grant (Code# PJ008148) from BioGreen21 Program, Rural Development Administration, Republic of Korea.