Severe Fever with Thrombocytopenia Syndrome (SFTS) is a newly emerging tick-borne disease caused by the SFTS virus (SFTSV), which belongs to the phlebovirus in the Bunyaviridae family. SFTSV is enveloped with a tripartite ambisense RNA genome. The L segment encodes the viral RNA-dependent RNA polymerase, the M segment encodes the two glycoproteins, Gc and Gn, and the S segment encodes the nucleoprotein (NP) and the nonstructural protein (NSs). NP participates in ribonucleoprotein (RNP) packaging and commonly detected early after infection, suggesting that the N protein is possible to be used as a target antigen for early diagnosis of SFTSV infection. In this study, we analyzed a highly immunogenic multi-epitope using GnGc and NP genes from a consensus sequence of SFTSV strain isolated from infected patients in Korea. The selected genes are constructed to the expression vector plasmid pJHL65 and the recombinant plasmid vector was transformed into the Δasd Δlon ΔcpxR Salmonella Typhimurium attenuated strain JOL912 and the expression of these antigens was verified by immunoblotting assay. We observed the significant levels of systemic IgG and mucosal IgM responses against the JOL912-derived antigen in the immunized BALB/c mice. The level of CD3+CD4+, CD3+CD8+ T lymphocyte subpopulation and TNF-α were also highly regulated in splenic T cells re-stimulated in vitro with NP and Gn/Gc multi-epitope selected antigens. Therefore, immunized mice with NP and Gn/GC multi-epitope recombinant proteins of attenuated Salmonella delivery system elicited T cell-related immune response, inducing an effective immune response. In conclusion, the attenuated Salmonella expressing NP-GnGc multi-epitopes could be a novel vaccine candidate against the SFTS virus.

Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe respiratory disease in humans, with a case fatality rate of approximately 35%, thus posing a considerable threat to public health. A lack of approved vaccines or antivirals currently constitutes a barrier for controlling disease outbreaks and spread. In this study, we generated a replication-defective recombinant vesicular stomatitis virus expressing the MERS-CoV spike (S) protein (VSV/MERS). Uncleaved and cleaved S proteins were detected in VSV/MERS by western blotting. And VSV/MERS specifically transduced cells expressing human dipeptidyl peptidase 4, a receptor for MERS-CoV. To investigate the immunogenicity of VSV/MERS, mice were immunized intramuscularly with VSV/MERS and alum adjuvant. MERS-CoV S-specific IgG was detected in serum samples from immunized mice. These antibodies inhibited MERS entry in vitro, suggesting a protective efficacy of VSV/MERS immunity. The data demonstrate that VSV/MERS has potent immunogenicity and could serve as a novel potential vaccine platform for MERS in dromedary camels and human.

This study examined the language choice for modality patterns to express the degree of probability specifically in email texts based on Systemic Functional Grammar (SFG) analytical framework. Participants were students enrolled in an English writing composition course at an online-based Korean university. They were required to write an email thanking their professor, in which they stated their future plans (definite or indefinite) with a degree of probability. The text analysis was compared with two groups of students' scores (high-scoring group n=40 and low-scoring group n=29), based on an assessment of the course assignment. After building up two learner corpora, UAM Corpus Tool version 3.0 was used to analyze the language choice closely using a modality system of the SFG framework. The high-scoring group showed more range and frequency in the use of modal verbs combined with a modal Adjunct or another modal expression. Explicit teaching on the importance of expressing the appropriate degree of probability using a range of modality devices, rather than relying heavily on the primary modality (choice of modal verbs) is highlighted as a pedagogical implication.

Acute vascular rejection has been known as a main barrier occurring in a xenograted tissue of alpha 1,3-galactosyltransferase knock-out (GalT KO) pig into a non-human primate (NHP). Adenosine which is a final metabolite following sequential hydrolysis of nucleotide by ecto-nucleotidases such as CD39 and CD73, act as a regulator of coagulation, and inflammation. Thus xenotransplantation of CD39 and CD73 expressing pig under the GalT KO background could lead to enhanced survival of recipient NHP. We constructed a human CD39 and CD73 expression cassette designed for endothelial cell-specific expression using porcine Icam2 promoter (pIcam2-hCD39/hCD73). We performed isolation of endothelial cells (pAEC) from aorta of 4 week-old GalT KO and membrane cofactor protein expressing pig (GalT-MCP/-MCP). We were able to verify that isolated cells were endothelial-like cells using immunofluorescence staining analysis with von Willebrand factor antibody, which is well known as an endothelial maker, and tubal formation assay. To find optimal condition for efficient transfection into pAEC, we performed transfection with GFP expression vector using four programs of nucleofection, M-003, U-023, W-023 and Y-022. We were able find that the program W-023 was optimal for pAEC with regard to viability and transfection efficiency by flow cytometry and fluorescent microscopy analyses. Finally, we were able to obtain GalT-MCP/-MCP/CD39/CD73 pAEC expressing CD39 and CD73 at levels of 33.3% and 26.8%, respectively. We suggested that pACE isolated from GalT-MCP/-MCP pig might be provided as a basic resource to understand biochemical and molecular mechanisms of the rejections and as an alternative donor cells to generate GalT-MCP/-MCP/CD39/CD73 pig expressing CD39 and CD73 at endothelial cells.

Double-stranded RNA (dsRNA) has been applied to control insect pests due to its induction of RNA interference (RNAi) of a specific target gene expression. Bacterial expression and formulation of dsRNA led to minimizing dsRNA degradation. In our studies overexpression of dsRNA specific to chymotrypsin, essential protease for the survival of the beet armyworm, Sopodoptera exigua, (SeCHY2) in Escherichia coli HT115 (DE3), was toxic to larvae after oral administration. In the other view, silencing of the transformer2 (tra-2) gene that seems to performance as the genetic modification generate female development in several dipteran species including the striped fruit fly, Baetrocera scutellata can used to development for male-only release technique to control of this pest.

Naegleria fowleri is pathogenic free-living amoeba leading to primary amoebic meningoencephalitis(PAM) in human and animals. The nfa1 gene cloned from N. fowleri is located on food-cup structure called pseudopodia and function an adherence of host target cells. To evaluate the effect of nfa1 vaccination against N. fowleri infection, we constructed retroviral vector(pQCXIN) expressing nfa1 gene. To determine the effect of vaccination and protective immunity in in vivo models, we measured the immunoglobulin levels, cytokine induction, and survival rate in mice infected with N. fowleri. Both levels of IgG and IgG subclass in DNA vaccinated mice were significantly elevated. The cytokine analysis show that DNA vaccinated mice induces production of IL-4 and IFN-γcytokines suggesting a Th1/Th2 mixed type immune response. The levels of nfa1 specific IgG antibody were maintained highly until 12 weeks post-vaccination in vaccinted mice. The nfa1 vaccinated mice using retroviral vector increased significantly survival rate(60%) after N. fowleri infection. Consequently, the nfa1 vaccination effectively induces protective immunity by upregulation of immune response in mice infected with N. fowleri. These results suggest that DNA vaccination using retroviral vector may be proper trial for treatment and prevention of PAM.

Transplantation is considered to be a very useful approach to improve human welfare and to prolong life-span. Heterologous organ transplantation using pig organs which are similar to human beings and easy to make mass-production has known as one of the alternatives. To ensure potential usage of the pig organ for transplantation application, it is essentially required to generate transgenic pig modifying immuno-related genes. Previously, we reported production of heterozygous α 1,3-galactosyltransferase (GalT) knock-out and human membrane cofactor protein (MCP) expressing pig (GalT-MCP/+), which is enforced for suppression of hyperacute and acute immunological rejection. In this study, we reported generation of homozygous pig (GalT-MCP/-MCP) by crossbreeding GalT-MCP/+ pigs. Two female founders gave birth to six of GalT-MCP/-MCP, and seven GalT-MCP/+ pigs. We performed quantitative real-time PCR, western blot, and flow cytometry analyses to confirm GalT and MCP expression. We showed that fibroblasts of the GalT-MCP/-MCP pig do not express GalT and its product Gal antigen, while efficiently express MCP. We also showed no expression of GalT, otherwise expression of MCP at heart, kidney, liver and pancreas of transgenic pig. Taken together, we suggest that the GalT-MCP/-MCP pig is a useful candidate to apply xenotransplantation study.

Mesenchymal stem cells (MSCs) have been considered an alternative source of neuronal lineage cells, which are difficult to isolate from brain and expand in vitro. Previous studies have reported that MSCs expressing Nestin (Nestin+ MSCs), a neuronal stem/progenitor cell marker, exhibit increased transcriptional levels of neural development-related genes, indicating that Nestin+ MSCs may exert potential with neurogenic differentiation. Accordingly, we investigated the effects of the presence of Nestin+ MSCs in bone-marrow-derived primary cells (BMPCs) on enhanced neurogenic differentiation of BMPCs by identifying the presence of Nestin+ MSCs in uncultured and cultured BMPCs. The percentage of Nestin+ MSCs in BMPCs was measured per passage by double staining with Nestin and CD90, an MSC marker. The efficiency of neurogenic differentiation was compared among passages, revealing the highest and lowest yields of Nestin+ MSCs. The presence of Nestin+ MSCs was identified in BMPCs before in vitro culture, and the highest and lowest percentages of Nestin+ MSCs in BMPCs was observed at the third (P3) and fifth passages (P5). Moreover, significantly the higher efficiency of differentiation into neurons, oligodendrocyte precursor cells and astrocytes was detected in BMPCs at P3, compared with P5. In conclusion, these results demonstrate that neurogenic differentiation can be enhanced by increasing the proportion of Nestin+ MSCs in cultured BMPCs.

폴리드나바이러스(polydnavirus: PDV)는 일부 내부기생봉에 공생하는 이중나선형 DNA 바이러스 분류군이다. Cotesia plutellae bracovirus (CpBV)는 프루텔고치벌(C. plutellae)에 공생하는 일종의 PDV이다. 프루텔고치벌은 어린 배추좀나방(Plutella xylostella) 유충에 기 생한다. 기생 초기에 발현하는 CpBV-ELP1 유전자는 혈구세포에 세포독성을 발휘하면서 기주의 세포성 면역을 억제하여 기생에 중요한 역할을 담당하고 있다. 본 연구는 이 유전자를 담배 식물에서 발현하여 해충에 대한 경구독성을 분석하는 데 목적을 두었다. 재조합 CpBV-ELP1 단백질 이 배큘로바이러스 발현시스템을 통해 합성되어 세포배양액에 분비되었다. 수거된 세포배양액은 일련의 단백질 분리과정(ammonium sulfate 단백질 분획, size exclusion 크로마토그래피, 이온교환 크로마토그래피)을 통해 CpBV-ELP1 단백질을 분리하는 데 이용되었다. 분리된 rCpBV-ELP1 단백질은 파밤나방(Spodoptera exigua) 혈구에 대한 뚜렷한 세포독성을 보였다. CpBV-ELP1은 파밤나방 5령충에 대해서 혈강 주입하여 살충력을 나타냈고, 엽침지법을 이용하여 경구독성을 갖고 있는 것을 확인하였다. CpBV-ELP1 유전자를 CaMV 35S 유전자 프로모터 와 opaline synthase 유전자 전사종결신호를 갖는 pBI121 벡터에 클로닝하여 Agrobacterium tumefaciens LBA4404 세균에 형질전환을 유도하였 다. 형질전환된 세균은 담배(Nicotiana tabacum Xanthi)잎에 감염하여 캘러스를 유도하게 하였고 이후 차세대(T1)를 확보하였다. T1 세대 담배 는 파밤나방에 대한 해충저항성을 갖고 있음을 확인하였다. 이러한 결과는 CpBV-ELP1 유전자가 형질전환작물을 통해 해충방제에 응용될 수 있 다는 것을 제시하고 있다.

In this paper, some popular intensity measures of earthquakes including magnitude, MMI, and PGA as well as their empirical relationships are briefly reviewed since they have been widely used without prudence by mass media, the public, and even the government when asking or expressing the seismic capacity of buildings. The basic concept of current seismic design is also presented in order to facilitate relevant discussions. It is emphasized that expressing the building seismic capacity simplistically in terms of seismological quantities or terminologies like magnitude and MMI is inherently irrational, may be misleading the stakeholders, and should be avoided. Alternative expressions, more rational and consistent with current seismic design philosophy and practice, are recommended.

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.

In all mammalian species, progesterone is essential in the preparation for and maintenance of pregnancy, if it occurs. Progesterone primes the endometrium for possible implantation and inhibits uterine contraction until birth. 20-alpha hydroxysteroid dehydrogenase (20α-HSD; EC.1.1.1.149) enzyme belongs to the family of aldo-keto reductases. 20α-HSD predominantly converts progesterone into its biologically inactive form 20α-hydroxyprogesterone (20α-OHP), and plays a crucial role in the termination of pregnancy and initiation of parturition. In addition, the activity of 20α-HSD during the luteal phase known to be inhibited by prolactin. In this study, we focused on the analysis of transgenic mice expressing EGFP under control of monkey 20α-HSD promotor in mice testis. The protein expression and localization were detected by Western blotting and Immunohistochemical analysis, respectively. 20α-HSD protein was detected at molecular weight of 37-kDa by Western blotting analysis and EGFP was found at 27-kDa in the testis of TG mice. Also EGFP and 20a-HSD protein expression on 1, 2, 4, 6 and 8 weeks after birth were assessed. Both of them were increased the expression level time-dependently. 20α-HSD were strongly expressed in seminiferous tubule from 1 week after birth as seen in Immunohistochemical analysis. However, EGFP was strongly expressed in the seminiferous epithelial cells. Then, we determined the expression of EGFP mRNA in mice testis. Using primers specific for mouse EGFP, mRNA expression levels were analyzed by RT-PCR. The EGFP molecular weights is 400bp, qRT-PCR results using EGFP primer, The Cq value of the ratio decreased as the age increased. On this basis, mRNA were increased the expression level time-dependently. In conclusion, these observations suggest that the 20α-HSD in testis could be play a pivotal role in the spermatogenesis.

Recently, RNA interference (RNAi) technology has been emerged as a potent tool for pest control strategy. Based on the previous studies on RNAi via leaf disc-mediated systemic delivery of dsRNA and in planta expression of hairpin RNA by agroinfiltration, the coatomer subunit alpha (COPA) gene has been found to be a crucial target for RNAi against Tetranychus urticae. In current study, transgenic plants of Arabidopsis thaliana expressing COPA hairpin RNA were generated by the floral dip method. Putative transgenic plants were screened by PCR and positive transformants were subjected to bioassay using age-synchronized and host-adapted T. urticae. T. urticae feeding on plants expressing dsRNA/siRNA showed more than 80% mortality as compared to the mites feeding on control plants at 6 days post-infestation. Our data shows that in planta expression of hairpin gene such as COPA may serve as an effective way for the control of this important pest in ornamental and economically important plants.

Conflicting accounts of environmentally-friendly motives exist (see Chan, 2001; Hartmann & Apaolaza-Ibanez, 2012; Haws et al., 2014; Johnstone & Tan, 2015; Rashid, 2009; Royne et al., 2013). Recent research has turned to identity-society explanations (see Park & Lee, 2016). This research furthers this inquiry and narrows the gap. To understand environmentally-friendly clothing options (EFCO) motives better, this study uses uniqueness theory, which posits that consumers adopt dress different from mass fashion simply because it is unpopular (Snyder & Fromkin, 1977; Tian et al., 2001). Accordingly, environmentally-friendly attitudes should have nothing to do with the environment, but with norms, conformity, pressure (Law et al., 2004), and uniqueness. Thus, the research questions consist of: 1) is need for uniqueness in dress related to EFCO purchase intentions? 2) If so, does uniqueness relate to other EFCO motives? A survey was administered (n=220), using existing scales, to an online consumer panel All scales exhibited sufficient reliability. Pearson-Product Moment Scores and ANOVAs were used to assess variable relationships. As predicted, concern for the environment and perceived individual impact on the environment were unrelated to need for uniqueness. There was a significant and positive relationship between need for uniqueness and each of: attitudes toward EFCO and social pressure to act green. This indicates that individuals feel social pressure from important others to adapt to consumer trends. However, the manner in which they adopt mass consumer movements, such as sustainability, may be in more unique ways and via unpopular choice, such as EFCO. Finally, an ANOVA indicated that those high in uniqueness were willing to pay substantially more for EFCO.

이중가닥 RNA (double-stranded RNA, dsRNA)는 표적 유전자의 발현을 억제하는 기능으로 해충방제에 응용되었다. 인테그린은 α와 β 단위체로 구성된 이량체 막 단백질이다. 진핵생명체에서 인테그린은 세포-세포 및 세포-세포외기질의 상호연결에 중요한 역할을 담당한다. 인 테그린 β 단위체 발현을 억제하는 특정한 dsRNA (= dsINT)는 해당 곤충에 뚜렷한 치사효과를 유발한다. 또한, dsINT를 발현시키는 형질전환 된 대장균도 해당 곤충에 뚜렷한 살충력을 가진다. 그러나 이 세균 살충제의 야외 적용을 위해서는 제형화 기술이 필요했다. 본 연구는 dsINT를 발현하는 재조합 세균을 동결 건조시켜 대상 곤충에 대해 살충효능을 검정하였다. 동결 건조된 세균은 파밤나방(Spodoptera exigua) 종령 유충에 높은 섭식독을 일으켰다. 파밤나방에 대해서 Bacillus thuringiensis 상용 살충제 처리는 불과 60%의 살충력을 보이는 반면, 동결 건조된 dsINT 발현 세균과 혼합 처리할 때 살충력은 크게 증가하였다. dsINT 발현 세균은 해당 인테그린 염기서열 유사성에 따라 차이를 보이는 해충 종에 선 택적 독성을 나타냈다. 이 결과는 인테그린에 특이적 dsRNA를 발현하는 세균이 동결 건조 제형화 조건하에서도 살충력을 유지한다는 것을 나타 냈다.

A large number of transgenic crop varieties expressing the Bt (Bacillus thuringiensis) insecticidal proteins have been commercialized in 13 countries since 1996. Although the use of these insect-resistant Bt crops can increase crop quality and yields, concerns remain about the potential negative effects of such crops on ecosystems. Transgenic soybean containing cry1Ac gene have been developed to control Lepidopteran pests of soybean and we aimed to investigate whether this soybean could affect non-target arthropods, which play a major role in ecological functions in agricultural ecosystems. In the present study, we first measured the levels of Cry1Ac protein in Bt soybean at different growth stages of soybean and then we compared the community structure of arthropods occurred in fields of transgenic and wild-type soybean. The levels of Cry1Ac protein in transgenic soybean leaves ranged from 252.9 to 604.5 μg g-1 DW. Multivariate analyses (PerMANOVA and NMDS) showed that the composition of the non-target arthropod community was affected by sampling date but not by soybean genotype. These results suggest that transgenic soybean expressing Cry1Ac protein may not adversely affect such non-target arthropod communities.

A subtropical insect species, Maruca vitrata, is invasive to temperate zones. Supercooling capacity of M. vitrata was confirmed in all developmental stages, in which egg exhibited the lowest supercooling point (SCP) at –22.5oC. However, low temperature lethality was observed at much higher temperatures than SCPs in all developmental stages. In addition, this nonfreezing injury increased with incubation time at rearing condition. Uncontrolled phenoloxidase activation appeared to cause the nonfreezing injury. Preexposure to a cool temperature before lethal low temperature significantly increased survival of M. vitrata. This cold hardening accompanied with increase of glycerol content in hemolymph. Transcriptome of M. vitrata under cond-hardening was assessed.

Polydnaviruses (PDVs) are a group of insect viruses and symbiotic to some endoparasitoid wasps. Genome analyses of different PDVs provide a number of genes putatively associated with alteration of host insect physiological processes. Especially, PDV gene products assist host wasp development by suppressing immune responses and delaying larval development of parasitized lepidopteran hosts. Thus, PDV genes can be applied to control insect pests by incorporating them into crop genomes. This talk illustrates two examples of PDV genes: CpBV-CST1 and CpBV-ELP1. CpBV-CST1 has been known to inhibit insect cysteine proteases to suppress immune and development, while CpBV-ELP1 exhibits a high cytotoxicity to insect cells. Transgenic tobaccos expressing CpBV-CST1 or CpBV-ELP1 were constructed by using Agrobacterium-mediated transgenic system and exhibited antixenosis against chewing and sucking insect pests on tobacco.

Xenotransplantation of pig islet regarded as a good alternative to allotransplantation. However, cellular death mediated by hypoxia-reoxygenation injury after transplantation disturb success of this technique. In the present study, we produce transgenic pig expressing human heme oxygenase 1 (HO1) genes to overcome cellular death for improving efficiency of islet xenotransplantation. Particularly, Korean miniature pig breed, Micro-Pig, was used in the present study. Somatic cell nuclear transfer (SCNT) technique was used to produce the HO1 transgenic pig. Six alive transgenic piglets were produced and all the transgenic pigs were founded to have transgene in their genomic DNA and the gene was expressed in all tested organs. Also, in vitro cultured fibroblasts derived from the HO1 transgenic pig showed low reactive oxygen species level, improved cell viability and reduced apoptosis level