Human interferon alpha 2b (hIFNα-2b) is an important immune regulator widely used in clinic, for the treatment of chronic hepatitis, hairy cell leukemia, chronic myelogenous leukemia and multiple myeloma, etc. The clinically used hIFNα-2b is generally produced by E. Coli, which lacks the post-translational O-glycosylation of naturally synthesized protein, and has a short serum half-life. In this study, we report the successful generation of transgenic chickens that produce hIFNα-2b in the egg white using a feline immunodeficiency virus (FIV)-based lentiviral vector. In preliminary in vitro study, the hIFNα-2b gene under the control of CMV promoter expressed as much as 2,650 ng/㎖ in CEF-LNC-hIFNα-2bW cell. A FIV vector packaged with vesicular stomatitis virus G glycoprotein (VSV-G) was injected underneath the blastoderm of freshly laid chicken eggs (stage X) to produce a hIFNα -2b transgenic chicken. Out of 187 injected eggs, 55 chicks were hatched after 21 days of incubation, and 27 of the G0 hatched chicks expressed the vector-encoded hIFNα-2b gene. The expression of recombinant hIFNα-2b in transgenic chickens constitutes an important step towards low-cost and full biological activity production of this protein drug in bioreactor. This work was supported by the Bio-industry Technology Development Program, Ministry of Agriculture, Food and Rural Affairs, Republic of Korea, and by a grant from the Next-Generation BioGreen 21 Program (No. PJ011178), Rural Development Administration, Republic of Korea.

In the present study, using a MoMLV-based retrovirus vector, we successfully generated a new transgenic chicken line expressing high levels of hEPO. A replication-defective Moloney murine leukemia virus (MoMLV)-based vectors packaged with vesicular stomatitis virus G glycoprotein (VSV-G) was injected beneath the blastoderm of non-incubated chicken embryos (stage X). One rooster was mated to wild-type hens to produce 748 G1 progeny. PCR analysis of blood samples from these progeny revealed that there were seven G1 transgenic offspring, corresponding to a 0.9% germline transmission rate. Subsequently, Southern blot analysis of the genomic DNA from three G1 transgenic chickens was carried out to verify the stable genomic integration and copy number of the transgene in the genome. Quantitative analyses of the blood samples taken from G1 transgenic chickens resulted in 4,150 ～ 10,823 IU/㎖ (34.6 ～ 90.2 ㎍/㎖) of hEPO in the blood. The biological activity of the recombinant hEPO in transgenic chicken serum was comparable to its commercially available counterpart. Red blood cell numbers were more than three-fold higher in the transgenic chickens compared to the non-transgenic chickens. Successful germline transmission of the transgene was also confirmed in G2 transgenic chicks produced from crossing G1 transgenic roosters with non-transgenic hens. We confirmed that 13 transgenic chicks of 45 G2 progeny, corresponding to a 28.9% germline transmission rate. These results will help establish a useful transgenic chicken model system for studies of embryonic development and for efficient production of transgenic chickens as bioreactors. This work was supported by the Bio-industry Technology Development Program, Ministry of Agriculture, Food and Rural Affairs, Republic of Korea, and by a grant from the Next-Generation BioGreen 21 Program (No. PJ011178), Rural Development Administration, Republic of Korea.

Hydroxyapatite (HA), which is an important calcium phosphate mineral, has been applied in orthopedics, dentistry, and many other fields depending upon its morphology. HA can be synthesized with different morphologies through controlling the synthesis method and several parameters. Here, we synthesize various morphologies of HA using two simple methods: hydrothermal combustion and solution combustion. The phase purity of the synthesized HA is confirmed using X-ray diffractometry. It demonstrates that pure phased hydroxyapatite can be synthesized using both methods. The morphology of the synthesized powder is examined using scanning electron microscopy. The effects of pH and temperature on the final powder are also investigated. At 140 oC, using the hydrothermal method, nano-micro HA rods with a hexagonal crystal structure can be synthesized, whereas using solution combustion method at 600 oC, a dense cubic morphology can be synthesized, which exhibits monoclinic crystal structures.

We quantified temporal and spatial changes in the habitat for fish populations, the distribution of mandarin fish(Siniperca scherzeri) and an introduced species, largemouth bass(Micropterus salmoides) in Lake Paro and inflowing streams. The number of fish species identified in Lake Paro and the tributary streams included 10 families, 24 species and 10 families 30 species, respectively. The dominant fish species in Lake Paro were Zacco platypus, Hemibarbus labeo, Squalidus gracilis majimae, S. scherzeri and Tridentiger brevispinis, Z. platypus, Z. koreanus, and S. gracilis majimae in the inflowing streams. Although the habitat segregation for S. scherzeri and M. salmoides occurs, these two species showed the use of the fishes of the family Gobiidae as an important prey item based on IRI analysis. S. scherzeri and M. salmoides preyed mainly on T. brevispinis(67.4 %) and R. brunneus(84.0 %), respectively. The species preyed on by S. scherzeri and M. salmoides were benthic fishes that inhabit shallow water depths around the lake and have little swimming ability.

Activated carbons (ACs) were prepared by activation of coal tar pitch (CTP) in the range of 700°C-1000°C for 1-4 h using potassium hydroxide (KOH) powder as the activation agent. The optimal activation conditions were determined to be a CTP/KOH ratio of 1:4, activation temperature of 900°C, and activation time of 3 h. The obtained ACs showed increased pore size distribution in the range of 1 to 2 nm and the highest specific capacitance of 122 F/g in a two-electrode system with an organic electrolyte, as measured by a charge-discharge method in the voltage range of 0-2.7 V. In order to improve the performance of the electric double-layer capacitor electrode, various mixtures of CTP and petroleum pitch (PP) were activated at the optimal activation conditions previously determined for CTP. Although the specific capacitance of AC electrodes prepared from CTP only and the mixtures of CTP and PP was not significantly different at a current density of 1 A/g, the AC electrodes from CTP and PP mixtures showed outstanding specific capacitance at higher current rates. In particular, CTP-PP61 (6:1 mixture) had the highest specific capacitance of 132 F/g, and the specific capacitance remained above 90% at a high current density of 3 A/g. It was found that the high specific capacitance could be attributed to the increased micro-pore volume of ACs with pore sizes from 1 to 2 nm, and the high power density could be attributed to the increased meso-pore volume.

In this work, we report in-situ observations of changes in catalyst morphology, and of growth termination of individual carbon nanotubes (CNTs), by complete loss of the catalyst particle attached to it. The observations strongly support the growth-termination mechanism of CNT forests or carpets by dynamic morphological evolution of catalyst particles induced by Ostwald ripening, and sub-surface diffusion. We show that in the tip-growth mode, as well as in the base-growth mode, the growth termination of CNT by dissolution of catalyst particles is plausible. This may allow the growth termination mechanism by evolution of catalyst morphology to be applicable to not only CNT forest growth, but also to other growth methods (for example, floating-catalyst chemical vapor deposition), which do not use any supporting layer or substrate beneath a catalyst layer.

This study was carried out to investigate the effects of stocking density on eating and ruminating behavior of Hanwoo steers (Bos taurus coreanae) in the finishing period. A total of 30 finishing Hanwoo steers (631.3 ± 11.4 kg, 25 months old) were allocated to one of four stocking density groups comprising 1, 2, 3, and 4 steers per 32 m2 pen [G1 (32 m2), G2 (16 m2), G3 (10.7 m2) and G4 (8 m2), respectively] in triplicate. Eating, rumination behaviors, as well as dry matter intake of steers were measured, and the results were subjected to analysis of variance with stocking density as the main effect. The results of eating behaviors over 48 hours are summarized as follows: Total intake was significantly (p<0.01) higher in G1, G2, and G3 compared to G4. Eating time was not different among the treatments, whereas ruminating time increased in the order of G1 > G2 > G3 > G4 (p<0.01). However, resting time and chewing time (sum of eating and ruminating) were not significantly different among the treatments. Number of boluses and number of total chews were highest in G1 (p<0.01), whereas number of chews per bolus was highest in G3 (p<0.01). Ruminating time per bolus as well as number of boluses per minute was not significantly different among the treatments. Number of defecations was higher in G1 and G2 animals compared to G3 and G4 animals (p<0.01). However, stocking density had no effect on drinking or urination. In conclusion, increasing stocking density (i.e. G4) per pen decreased voluntary intake, ruminating time, and total chewing number in the finishing period of Hanwoo steers. However, care must be taken in discussing stocking density in the present study as the space allowance per animal was satisfactory to meet the current animal welfare regulation in Korea and in Europe, although the beef production system in Korea is more intensive than in Europe.

Methanol as a carbon source in chemical vapor deposition (CVD) graphene has an advantage over methane and hydrogen in that we can avoid optimizing an etching reagent condition. Since methanol itself can easily decompose into hydrocarbon and water (an etching reagent) at high temperatures [1], the pressure and the temperature of methanol are the only parameters we have to handle. In this study, synthetic conditions for highly crystalline and large area graphene have been optimized by adjusting pressure and temperature; the effect of each parameter was analyzed systematically by Raman, scanning electron microscope, transmission electron microscope, atomic force microscope, four-point-probe measurement, and UV-Vis. Defect density of graphene, represented by D/G ratio in Raman, decreased with increasing temperature and decreasing pressure; it negatively affected electrical conductivity. From our process and various analyses, methanol CVD growth for graphene has been found to be a safe, cheap, easy, and simple method to produce high quality, large area, and continuous graphene films.

Chicken Insulin-like Growth Factor-1 (cIGF-1), one of the most important hormone for regulating physiological function includes body growth, muscle volume, bone density, chicken cell development and metabolism. In order to find in vitro Knokdown expression of cIGF-1, this study introduced tetracycline inducible RNA interference expression system (TetRNAi system). Tet system can inductively control high expression of extrinsic genes and expression of intrinsic genes. So it has advantages such as minimized physiological side-effects any cell and low cytotoxicity. RNAi system is proving to be a powerful experimental tool for inhibition of gene expression and post-transcriptional mechanism of gene silencing. RNAi is mediated by small interfering RNA (siRNA) consisting of 19- to 23- nucleotide double-stranded RNA duplexes that promote specific endonucleolytic cleavage of mRNA targets through an RNA-induced silencing. Then, this study RNAi-based gene knockdown can be achieved by retroviral-based expression systems. Stable integration of our inducible siRNA vector allowed the production of siRNA on doxycycline induction, followed by specific down regulation of chicken IGF-1 gene. Analyses of Real-time PCR to determine expression of the cIGF-1 gene showed successful from chicken embronic fibroblast (CEF) cells with the reduced rate of an approximately 92%. Our results demonstrate the successful regulation of cIGF-1 knockdown expression in CEF cells and support the application of an tetracycline inducible RNAi expression system in transgenic Mini chicken production. This research was supported by Bio-industry Technology Development Program, Ministry for Food, Agriculture, Forestry and Fisheries, Republic of Korea.

The leaf beetle, Chrysolina aurichalcea (Coleoptera: Chysomelidae), is a pest damaging plants of Compositae. In order to understand the genetic diversity and geographic variation we sequenced a portion of mitochondrial COI gene (658 bp) and complete nuclear internal transcribed spacer 2 (ITS2) of the species collected from seven Korean localities. A total of 17 haplotypes (CACOI01 ~ CACOI17), with the maximum sequence divergence of 3.04% (20 bp) were obtained from COI gene sequence, whereas 16 sequence types (ITS2CA01 ~ ITS2CA16), with the maximum sequence divergence of 2.013% (9 bp) were obtained from ITS2, indicating substantially larger sequence divergence in COI gene sequence. Phylogenetically, the COI gene provided two haplotype groups with a high nodal support (≥ 87%), whereas ITS2 provided one sequence type group with a high nodal support (≥ 92%). The result of COI gene may suggest the presence of historical biogeographic barriers that bolster genetic subdivision in the species. Different grouping pattern between COI gene and ITS2 sequences were interpreted in terms of recent dispersal, reflected in the ITS2 sequence. Finally, finding of unique haplotypes and sequence types only from Beakryeng-Islet population was interpreted as an intact remnant of ancient polymorphism. As more samples are analyzed using further hyper-variable marker, further fruitful inference on the geographic contour of the species might be available.

The leaf beetle, Chrysolina aurichalcea (Coleoptera: Chysomelidae), is a pest damaging plants of Compositae. In order to understand the genetic diversity and geographic variation of the species we sequenced a portion of mitochondrial COI gene (658 bp) and complete nuclear internal transcribed spacer 2 (ITS2) collected from seven Korean localities. A total of 18 haplotypes (BARCA01 ~ BARCA18), with the maximum sequence divergence of 3.04% (20 bp) were obtained from COI gene sequence, whereas 17 sequence types (ITS2CA01 ~ ITS2CA17), with the maximum sequence divergence of 2.013% (9 bp) were obtained from ITS2, indicating substantially larger sequence divergence in mitochondrial gene sequence. Phylogenetically, the mitochondrial DNA has shown several haplotypes formed independent groups with substantially high node support (≥ 90%), whereas no such grouping was evidenced for ITS2, indicating different behaviors of the two molecules. Such difference may reflect a diverse dynamics of the species such as biogeographic history, mating behaviors, and also possibly different mode of inheritance of the two molecules, but requires further scrutinized examination of the dataset. In terms of population genetic perspective, overall no population subdivision was detected from both molecules, except for locality 7 (Eocheong islet) from mitochondrial DNA. As more scrutinized analysis is performed, further fruitful inference on the geographic contour of the species might be available.

The seven-spotted lady beetle, Coccinella septempunctata (Coleoptera: Coccinellidae), known also as the seven-spot ladybird, is natural enemy for aphids and has a broad ecological range, living almost anywhere there are aphids for it to eat. In order to understand the genetic diversity and geographic variation of the species we sequenced a portion of mitochondrial COI gene (658 bp) and complete nuclear internal transcribed spacer 2 (ITS2) collected from nine Korean localities. A total of 21 haplotypes (CSCOI01 ~ CSCOI21), with the maximum sequence divergence of 4.56% (30 bp) were obtained from COI gene sequence (from 78 individuals), whereas 65 sequence types (CSITS201 ~ CSITS265), with the maximum sequence divergence of 2.06% (11 positions) were obtained from ITS2 (from 79 individuals), indicating substantially larger sequence divergence in mitochondrial gene sequence. Both COI gene and ITS2 shows the distribution pattern that only a few haplotypes or sequence types are widely distributed, whereas majority of them are highly restricted in one geographic location, even represented as a single individual. Unlikely the ITS2 sequence types the mitochondrial COI haplotypes evidenced the presence of two main phylogenetic groups, reciprocally monophyletic to each other. Geographically, these two groups are spread in all localities surveyed. Considering both COI gene and ITS2 sequence together, current our data may suggest the presence of ancestral polymorphism, rather than on-going speciation, but more scrutinized analysis will be performed soon. Due partially by the presence of both COI groups in all surveyed localities, the genetic diversity estimates of all localities are similar from the perspective of COI gene, but ITS data showed extremely lower genetic diversity of one islet locality, Anmyeon-do (locality 2; 0.002530 vs. 0.008054 ~ 0.012060). Analysis of gene flew estimates between localities indicates that most populations are highly interconnected to each other. However, one islet locality, Anmyeon-do (locality) has shown statistically significant distance from the remaining localities on the basis of only ITS2 data (FST = 0.19 ~ 0.34), requiring scrutinized phylogeographic inference on this population with expanded sampling. As more scrutinized analysis is performed, further fruitful inference on the geographic contour of the species might be available.

We report here the production of transgenic chickens that can regulate human erythropoietin (hEPO) gene expression. The glycoprotein hormone hEPO is an essential for viability and growth of the erythrocytic progenitors. Retrovirus vector system used in this study has two features including tetracycline-controllable promoter and woodchuck hepatitis virus posttranscriptional regulator element (WPRE). The former is for to reduce the possibility of physiological disturbance due to constitutional and unregulated expression of hEPO gene in the transgenic chicken. The latter is for maximum expression of the foreign gene when we turn-on the gene expression. A replication-defective Moloney murine leukemia virus (MoMLV)-based vectors packaged with vesicular stomatitis virus G glycoprotein (VSV-G) was injected beneath the blastoderm of non-incubated chicken embryos (stage X). Out of 325 injected eggs, 28 chicks hatched after 21 days of incubation and 16 hatched chicks were found to express the hEPO gene delivered by the vector. The biological activity of the recombinant hEPO in transgenic chicken serum was comparable to its commercially available counterpart. The recombinant hEPO in transgenic chicken serum had N- and O-linked carbohydrate simillar to that produced from in vitro cultured cells transformed with hEPO gene.