tasks that require nematode extraction and microscopic examination. To develop a more efficient detection method for Bursaphelenchus xylophilus, we first generated monoclonal antibodies (MAbs) specific to B. xylophilus. Among 2,304 hybridoma fusions screened, a hybridoma clone named 3-2A7-2H5 recognized a single protein from B. xylophilus specifically. We finally selected the MAb clone 3-2A7-2H5-D9-F10 (D9-F10) for further studies. To identify the antigenic target of MAb-D9-F10, we analyzed proteins in spots, fractions or bands via nano liquid chromatography electrospray ionization quadrupole ion trap mass spectrometry (nano-LC-ESI-Q-IT-MS). Peptides of galactose-binding lectin-1 of B. xylophilus (Bx-LEC-1) were commonly detected in several proteomic analyses, demonstrating that this LEC-1 is the antigenic target of MAb-D9-F10. The localization of MAb-D9-F10 immunoreactivities at the area of the median bulb and esophageal glands suggested that the Bx-LEC-1 may be involved in food perception and digestion. The Bx-LEC-1 has two non-identical galactose-binding lectin domains important for carbohydrate binding. The affinity of the Bx-LEC-1 to D-(+)-raffinose and N-acetyllactosamine were much higher than that to L-(+)-rhamnose. Based on this combination of evidences, MAb-D9-F10 is the first identified molecular biomarker specific to the Bx-LEC-1.

Three acetylcholinesterases (AChEs) were identified from the pinewood nematode, Bursaphelenchus xylophilus. Sequence comparison with known AChEs in conjunction with three-dimensional structure analysis suggested that all BxAChEs share typical characteristics of AChE at the major catalytic structures. BgAChE3 was most predominantly transcribed and then followed by AChE1 and AChE2. Immunohistochemistry using anti-BxAChEs antibodies revealed that BxAChE1 is most widely distributed whereas BxAChE2 exhibits more localized distribution in neuronal tissues. BxAChE3 was detected from entire body together with some limited tissues, including mouth parts and alimentary lining, and determined to be the only soluble AChE, suggesting its localization in hemolymph or/and extracellular space. Kinetic analysis of in vitro expressed BxAChEs revealed that BxAChE1 has the highest substrate specificity whereas BxAChE2 has the highest catalytic efficiency with BxAChE3 having the lowest catalytic efficiency. Interestingly, presence of BxAChE3 in the pool of BxAChEs significantly reduced the inhibition of BxAChE1 and BxAChE2 by inhibitors. Knockout of BxAChE3 by RNAi significantly increased the toxicity of nematicides, suggesting the protective role of BxAChE3 against these toxicants. Based on several features, including tissue distribution, expression level, substrate kinetics and inhibition property, it appeared that BxAChE1 is the major AChE with the function of postsynaptic transmission whereas BxAChE3 has been evolved to acquire the function of chemical defense, perhaps intrinsically against secondary toxic compounds from host pine trees, such as α-pinene and limonene. BxAChE2 appears to play a role in post-synaptic transmission in specialized neurons but its detailed physiological function still remains to be elucidated.

The Classical Swine Fever Virus (CSFV) is a member of the Pestivirus genus of the Flaviviridae. The genome of CSFV is a positive single-stranded RNA molecule 12.3 kb and contains a single large open reading frame (ORF). The polyprotein composed of eight nonstructural and four structural proteins (nucleocapsid protein C and three envelope glycoprotein E0, E1 and E2). E2, the most immunogenic of the CSFV glycoproteins, induces a protective immune response in swine. To determine the characteristics of the CSFV, LOM strain, we investigated the nucleotide sequence of the glycoprotein E0, E1 and E2. Comparison of the LOM with the other strains revealed nucleotide sequence identity ranging from 97 to 98%. Expression of the glycoprotein E2 was identified by SDS-PAGE and Western blot analysis using anti-CSFV E2 monoclonal antibodies in Sf21 cells. The expression levels of glycoprotein E2 were observed from day 3 and 5 days maximum. In addition, its expression efficiency by media and cell line was investigated. The result showed that High-Five cells and Grace’s insect media for Sf21 were the best conditions for the expression of the glycoprotein E2.

Aujeszky’s disease (AD), also called pseudorabies, is an infectious viral disease, caused by an alpha herpes virus and has domestic and wild pigs, as well as a wide range of domestic and wild animals, as the natural host. AD affects many countries and regions in the world, causing important economic losses, mainly due to international trade restrictions. In this study, to determine the characteristics of the Aujeszky’s disease virus (ADV), NYJ strain, which was isolated from the serum of an infected pig in 1987, we investigated the nucleotide sequence and expression of the glycoproteins gB, gC, and gD using the bBpGOZA system. We found that the glycoproteins gB, gC, and gD of NYJ consisted of 2751 bp, 1443 bp, and 1203 bp, respectively. Comparison of the NYJ with the other strains revealed nucleotide sequence identity ranging from 91.tito 99.0%. To better understand the genetic relationships between other strains, phylogenetic analyses were performed. The NYJ strain was formed a distinct branch with high bootstrap support. The expression of glycoprotein gD in insect cells was characterized by SDS-PAGE and Western blotting with an anti-ADV polyclonal antibody. Glycoprotein gD of approximately 45 kDa was detected. The results of this study have implications for both the taxonomy of ADV and vaccine development.

Bee venom contains a variety of toxic enzymes and peptides. One of the major components of bumblebee venom is bombolitin, which is the most abundant venom constituent and biologically similar to melittin. Here, we first show the molecular cloning and antimicrobial activity of the venom bombolitin from the bumblebee Bombus ignitus. The B. ignitus venom bombolitin gene consists of 2 exons, encoding 56 amino acid residues. The bombolitin purified from B. ignitus venom was the 2104 Da mature peptide with 18 amino acid residues, which are created by cleavage of the probombolitin domain between Ala38 and Leu39. We examined the pattern of bombolitin expression to confirm that it is a component of bumblebee venom. B. igniutus venom bombolitin exhibits venom gland-specific expression. We also investigated the venom bombolitin for antimicrobial properties against bacteria and fungi. The venom bombolitin showed high antibacterial activity against both Gram-negative and Gram-positive bacteria. Most interestingly, the venom bombolitin showed high antifungal activity against Fulvia falva, a leaf mold, and Alternaria radicia, a black rot. These antimicrobial profiles of B. ignitus venom bombolitin reported herein will be useful in the application for potential antimicrobial agents.

The pinewood nematode (PWN, Bursaphelenchus xylophilus) is known as a virulent factor of the pine wilt disease, transmitted to pinewoods by the pine sawyer beetle, Monochamus alternatus. It is very hard to discriminate B. xylophilus from B. mucronatus because these Bursaphelenchus species are genetically and biochemically very close. Therefore, it has been necessary to detect PWN-infected trees for the prevention of pine wilt disease transmission in a short time. We developed polyclonal antibodies against B. xylophilus in BalbC mice and primarily screened with ELISA. Positive clones releasing polyclonal antisera revealed B. xylophilus-specific immuno-reactivity, which were at least two times higher than that of B. mucronatus. Two clones, D9-F10 and 1F3, were finally selected and exhibited specific immuno-reactivity for B. xylophilus, not for B. mucronatus in Western blot analysis. D9-F10 clone was reactive with a 43-kDa whereas 1F3 clone with two proteins, 40- and 45-kDa. Their isotypes against mouse Ig family were identical, kappa-light chain. These results suggest that these monoclonal antibodies can be useful for the development of diagnostic kit for the pine wilt disease.

To determine the characteristics of the Korean porcine reproductive and respiratory syndrome virus (PRRSV), CA, which was isolated from the serum of an infected pig in 2006, we investigated the nucleotide sequence and expression of the structural ORFs (ORFs 2 to 7) using the bApGOZA system. We found that the structural ORFs 2 to 7 of CA consisted of 3188 nucleotides that were the same as those formed from VR-2332. Comparison of the CA with the other strains revealed nucleotide sequence identity ranging from 89.8 to 99.5%. To better understand the genetic relationships between other strains, phylogenetic analyses were performed. The CA strain was closely related to the other North American genotype strains but formed a distinct branch with high bootstrap support. Additionally, expression levels of the PRRSV proteins in Sf21 cells were strong or partially weak. The results of this study have implications for both the taxonomy of PRRSV and vaccine development.

The porcine reproductive and respiratory syndrome virus (PRRSV) has six structural proteins which encoded by ORFs 2 to 7 are designated as GP2, 3, 4, 5, M and N, repectively. In this study, we determined the expression of each protein using novel transfer vector, pBmKSK4 which has the polyhedrin promoter of BmNPV and 6xHis tag. The recombinant transfer vector was co-transfected into Bm5 cells along with bBpGOZA DNA. Recombinant virus was purified by plaque assay and amplified in Bm5 cells. Expression of each protein was identified by SDS-PAGE and Western blot analysis using anti-6xHis monoclonal antibody. The expression levels of the structural proteins in Bm5 cells were stronger than the expression system using pBacPAK9 transfer vector in Sf21 cells. As expected, GP5 was expressed at low levels from its structural properties and its toxicity for cells. In addition, each recombinant protein was purified using Ni-NTA spin columns. The ability to produce each protein in the baculovirus system indicates that these could be major candidates for the development of a vaccine against PRRSV.

The pine sawyer beetle, Monochamus alternatus, transmits the pinewood nematode (PWN, Bursaphelenchus xylophilus), causing the pine wilt disease (PWD), which gives rise to enormously economic as well as forest damage. However, PWN has been identified as a pathogen of PWD, it is very difficult to discriminate B. xylophilus from B. mucronatus in a short time, which are genetically and morphologically very similar. Therefore, it has been necessary to detect and eliminate PWN-infected trees in the forest area for the prevention of PWD transmission. Up to date, there is no report on biomarkers such as DNA and protein for the diagnosis of B. xylophilus. In this study, we produced a B. xylophilus monoclonal antiserum (D9-F10) from BalbC mice and screened its specificity with various proteins extracts. Western blot analysis revealed that the D9-F10 is only reactive with B. xylophilus protein extract among other tested protein extracts, indicating that the D9-F10 is specific for a B. xylophilus protein. Furthermore, two-dimensional electrophoresis showed the D9-F10 detects a very highly acidic protein, pI≒3.5. These results suggest that the D9-F10 monoclonal antibody is useful for the development of a diagnostic kit for the pine wilt disease.

Insect nicotinic acetylcholine receptors (nAChRs) are targets for insecticides. Despite the importance of the nAChR as a major target for insecticide action, modulators of nAChRs in insects remain unidentified. Here we describe the cloning and identification of a nAChR modulator gene in an insect. This gene was isolated by searching the firefly Pyrocoelia rufa cDNA library, and the geneitself encodes a protein 120 amino acids in length, named Pr-lynx1. Pr-lynx1 shares all the features, including a cysteine-rich consensus motif and common gene structure, of the Ly-6/neurotoxin superfamily. The recombinant Pr-lynx1, which is expressed as a 12-kDa polypeptide in baculovirus-infected insect Sf9 cells, is normally present at the cell surface asa GPI-anchored protein. Northern and Western blot analyses revealed that Pr-lynx1 is expressed in various tissues, such as the ganglion, brain, mandibular muscle, proventriculus, leg muscle, and epidermis. This expression pattern is similar to the distribution of nAChRs as assayed by α3 nAChR immunoreactivity. Co-expression of Pr-lynx1 in Xenopus oocytes expressing α3β4 nAChRs results in an increase in acetylcholine-evoked macroscopic currents, indicating a functional role of Pr-lynx1 as a protein modulator for nAChRs. This study on Pr-lynx1 is the first report of a modulator of nAChRs in an insect species.

The pinewood nematode (PWN, Bursaphelenchus xylophilus) causes the pine wilt disease, transmitted to pinewoods by the pine sawyer beetle, Monochamus alternatus. It is very difficult to discriminate B. xylophilus from B. mucronatus. Therefore, it has been necessary to detect PWN-infected trees for the prevention of pine wilt disease transmission in a short time. The development of biomarkers such as DNA and protein is important for diagnosis of B. xylophilus. However, there have been no reports regarding biomarker identifications of B. xylophilus. In this study, polyclonal antisera were raised against whole proteins of B. xylophilus in BalbC mice and were primarily screened with ELISA. Twenty five among over 500 cell lines releasing polyclonal antisera revealed B. xylophilus-specific immuno-reactivity, which was at least three times higher than that of B. mucronatus. Three cell lines among them were secreting monoclonal antibody through further screening. These cell lines only detect about a 33-kDa protein in B. xylophilus in the western blot. These results suggest that these monoclonal antibodies will be useful for the development of diagnostic kit for the pine wilt disease.

본 연구에서는 확산에 의해 발생하는 시멘트 방벽의 변질 과정을 반응성용질이동 모델링을 통해 장기간 동안 예측하고자 하였다. 모델링 결과 50,000년 후 시멘트의 변질은 30cm까지 진행되었다. pH는 13.0에서 11.86까지 감소하였으며 이는 알칼리 이온의 감소, 포틀랜다이트(portlandite)와 CSH (Calcium Silicale Hydrate)광물의 용해/침전반응에 의해 결정되고 있었다. 공극률 또한 portlandite와 의 용해에 의해 가장 큰 영향을 받고 있었으며 최고 약 0.3 점도 증가하였다. 우라늄의 용해도 역시 증가하고 있었으며 이는 pe의 증가에 기인하고 있었다. 본 연구 결과는 장기간의 시멘트 변질이 pH, pe, 공극률을 변화시킴으로서 핵종의 이동을 증가시킬 수 있음을 보여주고 있다.

본 연구에서는 핵폐기물 매립장의 인공 방벽으로 사용되는 시멘트 물질들과 주변 지하수 반응 결과로 형성되는 강알칼리성 지하수와 주변 암과의 반응을 통해 변화되는 지하수 특성을 지구화학 모델링을 통해 예측하고자 하였다. 연구 결과 시멘트 수화반응을 통해서 pH는 13.3를 나타내었으며 이때 생성되는 광물들은 Brucite, Katoite, Calcium Silicate Hydrate(CSH 1.1), Ettringite, Hematitie, Portlandite였다. 이들 광물들과 경주 지역에서 채취된 지하수의 반응 모델링에서는 지하수의 pH가 12.4로 예측되었다. 이러한 강알칼리성 지하수와 주변 화강암과의 반응은 년 동안 반응속도 모델링을 통해 모사하였다. 그 결과 지하수의 최종 pH는 11.2였으며 pH는 규산염 광물과 CSH 광물들의 용해 침전에 의해 조절되고 있었다. 또한 지하수 수질도 이들 광물들과 점토광물 및 산화광물들의 용해 침전에 의해 결정되고 있었다. 본 연구 결과는 장기간 동안의 강알칼리성 지하수와 주변 암과의 반응 모델링을 통해 지구화학 및 수질 변화를 예측함으로서 인공 방벽의 안정성 평가에 기여할 수 있을 것으로 판단된다.

원형질체 융합에 의한 화합성 및 불화합성 종간 체세포잡종을 얻었다. 화합성 종간인 Pleurotus ostreatus 와 P. florida 의 융합체는 이질핵체 (heterokaryon) 를 형성하였고, 불화합성 종간인 P. cornucopiae + P. florida , P. ostreatus + Ganoderma applanatum, P. florida + Ganoderma lucidum, 그리고 P. ostreatus + Flammulina velutipes 는 합핵체(synkaryon) 를 형성하였다. 이질이핵체는 동일한 양상의 자실체를 형성하는데 비해 합핵체는 유사분열상의 꺽쇠연결체 형성, 한쪽 친과 유사한 자실체 형성, 비정상적 유전형질 분리 및 유전자재조합 현상을 나타내었다. 화합성 및 불화합성 계통간 융합체의 RAPD 분석결과 화합성 종간 융합체는 동일한 DNA 패턴을 나타내었고, 불화합성 종간 융합체는 한쪽 친과 유사한 DNA 양상이면서 비양친 DNA 밴드도 형성하였다. 합핵체의 패턴은 microgenome insertion type 과 macrogenome insertion type 으로 구분되었다. 합핵체의 자실체 발생은 융합 모균주 양친의 자가임성에 의존하는데 이는 느타리의 동형핵체 자가임성과 유사한 양상이었고, 교배형 전환과 관련이 있는 것으로 사료된다. 여기서는 이러한 관점에서 논할 것이다.

골반저근은 골반기관을 지지하는 기능을 가지고 있으며 요자제를 유지하는 여성의 주요 하부조직이다. 골반저근의 약화는 복압성 요실금의 원인이 되는데, 이러한 골반저근의 기능 정도는 복압성 요실금의 병증정도를 평가하는 지표로 사용될 수 있다. 이에 본 연구에서는 골반저근의 수축 압력을 측정하여 복압성 요실금의 병적 진행정도를 정량적으로 진단할 수 있는 요실금 진단 알고리즘을 제안하였다. 이를 위하여 골반저근의 수축압력 정보를 측정할 수 있는 시스템을 제작하였으며, 측정된 데이터의 특징 분석을 위한 측정 프로토콜을 제안하였다. 복압성 요실금 환자로부터 획득한 데이터를 이용하여 5개의 진단 파라미터를 추출하였으며, 이를 이용한 진단 알고리즘을 구현하였다. 임상시험을 통하여 진단 알고리즘의 정확성을 평가한 결과 80%의 정확성을 보였으며, 20%의 위양성 진단 결과를 보였다. 반면에 위음성 진단 결과는 확인되지 않았다. 본 연구에서 제안한 요실금 진단 알고리즘은 복압성 요실금의 병적 진행 정도를 정량적으로 진단할 수 있으며, 요실금 진단 시스템 개발에 활용될 수 있을 것으로 판단된다.

Acute myocardial infarction (AMI) as an acute form and a catastrophic manifestation of coronary artery disease (CAD) is the major cause of morbidity and mortality worldwide. The prevalence and incidence of myocardial infarction can be influenced by demographic, biological and psychosocial factors, and it is rare in childhood and adolescent years. Contrary to the elderly, where cardiac attack is mainly caused by CAD, exercise-induced cardiac attack is relatively more prevalent in a young adult. We report here on a case of ST-segment elevation myocardial infarction (STEMI) in a young adult during vigorous physical activity and we review the relevant literatures.

This study was conducted to investigate the effects of rooting promoters on rooting and seedling quality in Buddleja davidii ‘Empire Blue’, Hydrangea paniculata ‘Magical Candle’, Hydrangea arborescence ‘Annabelle’, Stephanandra incisa semi-hardwood cuttings. IAA and IBA (500, 1000, or 1500 mg･L-1 soaking for 30 minutes and 3,000 mg･L-1 dipping for 10 seconds) or RootoneTM were used as rooting promoter. In B. davidii ‘Empire Blue’, IBA 1,500 mg･L-1 soaking reduced rooting percentage than control, but the effect of rooting promoters was generally insignificant. Number of leaves and shoot length were reduced in RootoneTM and IBA 1,500, 3,000 mg･L-1 treatments. IAA 1,000~3,000 mg･L-1 and IBA 500 mg･L-1 treatments showed better mat formation than control. The rooting ratio of H. paniculata ‘Magical Candle’ increased to 66.7% in IBA 1,500 mg･L-1 soaking treatment compared with 26.4% in the control. The mat formation was higher in RootoneTM, IAA and IBA treatments compared to control, and root length showed longer tendency in IAA and IBA except RootoneTM. The rooting ratio of H. arborescence ‘Annabelle’ was highest at 70.8% in the control and auxin treatments showed negative effect on the rooting percentage except IAA 1,000 mg･L-1 soaking treatment. The mat formation was very poor in IBA 1,000 and 1,500 mg･L-1 soaking at 1.1 and 1.3, respectively. S. incisa showed high rooting percentage at RootoneTM (80%) and IAA 1,500 mg･L-1 (80%) treatments compared to control (51.7%). Root length and dry weight were high in IAA 1,500 mg･L-1 soaking treatment. As a result, B. davidii ‘Empire Blue’ and H. arborescence ‘Annabelle’ did not need rooting promoter for stem cuttings, and IBA 1,500 mg･L-1, IAA 1,500 mg･L-1 soaking treaments could be used to improve rooting and seedling quality of H. paniculata ‘Magical Candle’ and S. incisa semi-hardwood cuttings, respectively.

Background : Angelica dahurica Bentham et Hooker, Angelica gigas Nakai, Ostericum koreanum Maximowicz and Peucedanum japonicum Thumberg　are a major medicinal plant in north Geungbuk province. Using medicinal plants are impotant it`s ingredient. Dry condition and stroage method are not standard manual. The ingredient variation of dry condition and stroage method were not researched. Methods and Results : Using plant material were cutivated on Gyongsangbukdo Bonghwa area. It were studied ingredient variation after dry and storage condition by HPLC methods. Major ingredient of Angelica gigas Nakai are decurusin, decurusinangelate. Heated air bulk dry get more decursin than natuarl dry and decurusinangelate of natural bulk dry was higher than heated air bulk dry. Major ingredient of Ostericum koreanum Maximowicz are imperatorin and isoimperatorin.. Imperatorin of Ostericum koreanum was highest peak on 50℃ heated-air dry after plastic bag sorage and isoimperatorin was highest peak on 40℃ heated-air dry after mountain cultivation. Imperatorin is a major ingredient Angelica dahurica Bentham et Hooker. Heated air bulk dry get more decursin and decursinangelate than natuarl dry and small heated-air dry. Peucedanol-7o_glucoside is a major ingredient Peucedanum japonicum Thumberg. Natural bulk dry get more peucedanol-7o_glucoside than heated-air bulk dry. Conclusion : Ingredient of Angelica dahurica Bentham et Hooker, Angelica gigas Nakai, Ostericum koreanum Maximowicz are different under various cutivation, drying method, storage. Diffent Ingedients of Angelica gigas Nakai, Ostericum koreanum Maximowicz were not accord it’s optical conditon.

Background : Malonyl ginsenoside content of the Panax ginseng is known to account for 35% to 60% of total ginsenosides content. However, its distribution by ginseng part has not been studied. In this study, four kinds of malonyl ginsenosides were compared in Korean white ginseng part using the purified malonyl ginsenoside standards in our laboratory. Methods and Results : White ginseng was prepared by the air drying (50℃, 48h) or freeze drying (-70℃, 48h) methods form 4-year-old ginseng. Malonyl ginsenoside content in total ginsenosides were similar in air dried and freeze dried white ginseng, 58% and 62%, respectively. Therefore, malonyl ginsenoside contents in main, lateral, and fine root, and in the main root without skin and skin of main root prepared by freeze dried method were compared. Malonyl ginsenosides (m-Rb1, m-Rb2, m-Rc and m-Rd) and total ginsenosides (Rg1, Re, Rf, Rb1, Rc, Rb2, Rb3, Rd, m-Rb1, m-Rb2, m-Rc and m-Rd) were 6.75 and 14.15 mg/g in main root, 14.15 and 26.35 mg/g in lateral root, 46.95 and 84.15 mg/g in fine root. Malonyl ginsenoside contents in skin of main root was 20.08 mg/g, while its contents of the main root without skin was 2.58 mg/g. Conclusion : As a result, the parts each air drying the sample was confirmed that the ratio of the distribution of malonyl ginsenoside (main root : lateral : fine root = 18.7 : 11.1 : 16.2), and distribution ratio of main root, skin of main root, lateral, skin of lateral was found to be (12.2: 14.6 14.3: 3.7). Malonyl ginsenoside content was the highest in fine root, compared to the main or lateral root. Malonyl ginsenoside contents in skin of root was higher than those of the main root without skin. These results is expected to help establish an efficient extraction and standardization. Malonyl ginsenoside analysis of White ginseng using HPLC expects that the standardization process can be established.

Background : Usually, planting and harvest time is around the middle of April. A labor shortage deu to her planting time as harvest time and some farmers were late planting culture. After planting until take rooting It will take more than one month when late planting growth is bad. Seedling cultivation, just taking rooting is agricultural growth promotion. Methods and Results : So did design the optimum nursery days and number of plants. Nursery days were 30 days and 45 days and number of plants 1, 3, 5 plants. Plant length was longer in less number of plants while Root length was longer in more number of plants of Temporary planting seedling, Leaf number was a big increase but number of plants was not a big shift. In post-havest survey, plant length was longer in 3 number of stem and root length was longer in one number of stem. 45 days and 3 number of stem seedling was the most good in number, weight and length of tuberous root. Yield of 45 days and 3 number of stem seedling was higher 7% than 1 number of stem seedling and higher 5% than 5 number of stem seedling. 45 days and 3 number of stem seedling was 1,789 thousand per 10a in income was the most. Conclusion : Result of this study, for rasing healthy seeding nursery days 45 days and number of plants 3 plant