The gene (2,304-bp) encoding a novel xylanolytic enzyme (XylD) with a catalytic domain, which is 70% identical to that of Cellulomonas flavigena DSM 20109 GH6 β-1,4-cellobiohydrolase, was identified from an earthworm (Eisenia fetida)-symbiotic bacterium, Cellulosimicrobium sp. strain HY-13. The enzyme consisted of an N-terminal catalytic GH6-like domain, a fibronectin type 3 (Fn3) domain, and a C-terminal carbohydrate-binding module 2 (CBM 2). XylDΔFn3-CBM 2 displayed high transferase activity (788.3 IU mg-1) toward p-nitrophenyl (PNP) cellobioside, but did not degrade xylobiose, glucose-based materials, or other PNP-sugar derivatives. Birchwood xylan was degraded by XylDΔFn3-CBM 2 to xylobiose (59.2%) and xylotriose (40.8%). The transglycosylation activity of the enzyme, which enabled the formation of xylobiose (33.6%) and xylotriose (66.4%) from the hydrolysis of xylotriose, indicates that it is not an inverting enzyme but a retaining enzyme. The endo-β-1,4-xylanase activity of XylDΔFn3-CBM 2 increased significantly by approximately 2.0-fold in the presence of 50 mM xylobiose.

The aim of this study was to develop Prevotella intermedia ATCC 49046-specific PCR primers designed based on the nucleotide sequence of a DNA probe Pig28. The strainspecificity of the PCR primers, Pig28-F1/Pig28-R1, was confirmed with 9 strains of P. intermedia and 25 strains (15 species) of Prevotella species. The detection limit of the PCR primers was 2 pg of the purified genomic DNA of P. intermedia ATCC 49046. These PCR primers were found to be useful for identifying P. intermedia ATCC 49046, particularly for determining the authenticity of the strain.

Canine parvovirus (CPV) type 2a (CPV-2a) has recently been identified as the main genotype circulating in the dog population in South Korea. Although CPV vaccines protect domestic dogs from CPV-2 infection, the efficacy of commercial live or inactivated CPV vaccines against CPV-2a has not been reevaluated. In this study, dogs were immunized with one of 7 commercial CPV vaccines (4 modified live and 3 inactivated vaccines) followed by challenge with CPV-2a strain, KV0901 that had been isolated from naturally infected dog in 2009. All dogs vaccinated twice with 4 commercial modified live CPV vaccines were seroconverted (geometric mean HI titer > 190.2) and most of dogs were completely protected against virulent CPV-2a strain infection. The dogs inoculated with 3 commercial inactivated CPV vaccines were also seroconverted and showed a slight loss of appetite and light diarrhea for 4 days after challenge and returned to normal at 5 days post challenge. However, the non-vaccinated dogs revealed the typical clinical signs of CPV infection including haemorrhgic diarrhea. In conclusion, the 4 live CPV vaccines licensed in Korea cross-protected dogs against virulent challenge with CPV-2a and are applicalble to pet dogs for the prevention of CPV infection.

Single basidiospores were isolated from a Pleurotus ostreatus strain collected in Boryeong, Korea showing gray and semispherical in color and shape of pileus. Mycelial growth and several other characters were good to be a breeding material. Mating experiments were performed using 23 monokaryons to provide mating compatibility data. Pairings were performed in 90 mm petri dishes on PDA. They were allowed to grow at 25℃ until two fronts of the advancing mycelia met and developed a conspicuous contact zone. The contact zone and the outer edges of paired colonies on each plate were examined for clamp connections and the crosses were considered positive when clamp connections were found at the growing margin of either side of the interacting monokaryons. The collected strain resulted in tetrapolar incompatibility system in intrastrain crosses. Compatible matings could be visibly distinguished from incompatible ones by the rapid growth and gross morphology. The dikaryotic colony was also confirmed microscopically by the presence of clamp connections from incompatible pairings. The four classes of tester strains representing the four incompatibility types were chosen from intrastrain mating tests as expected. The mating tests with the tester strain were performed for the determination of the rest monokaryons. As a result of these tests it was found that the strain share the same A and B incompatibility factors showing tetrapolar incompatibility system.

Agaricus bisporus is a nutritious edible fungus that is cultivated worldwide. Because of its secondarily homothallic life cycle, 90% or more of the basidia lining the gills at fruit bodies are predominantly two-spored, with each spore receiving two of the four meiotic nuclei. Such homothallic system results in scarcity of monokaryotic basidiospores in its progeny but the availability of uninucleate self-sterile homokaryons is a pre-requisite for producing hybrids in breeding program. This experiment screened 64 single spore isolates collected from CN0210-021, which is white button mushroom cultivated in Chungnam area. They were germinated on PAD meduium at 25℃. After 30 days of incubation colony diameter and morphology type of each isolates was determined. Based on growth rate, 5 strains showing very slow growing morphology than that of parental heterokaryotic strain were selected. Out of 5 slow growing isolates, 2 of them fruited and the rest 3 isolates did not fruit, which were considered as putative homokaryons. These data will provide the outcrossing sources that could be used in commercial breeding programs later. However, in the identification of homokaryotic strains, there is no direct morphological distinguishing features such as a clamp connection that would permit the reliable sorting between homokaryons and heterokaryons. Traditional method to verify a homokaryon thorough the fruiting trial is also time-consuming and not perfect. Therefore, to accelerate the breeding of Agaricus bisporus, quick and reliable methods such as molecular markers to identify the infrequent homokaryons from heterokaryons are needed to be carried out.

The aim of this study was to determine the immediate effects of single treatment of strain-counter strain (SCS) on pressure pain threshold (PPT) and muscle activity during scapular plane abduction with 3% body weight load. Fifteen asymptomatic male adults with upper trapezius latent trigger point (LTrP) (PPT<2.9 ) participated in this study. Pressure algometer was used to measure PPT and surface electromyography was used to record upper, middle arid lower trapezius, serratus anterior, infraspinatus and middle deltoid muscle activity and relative ratio during scapular plane abduction between pre- and post-intervention. There was a significant increase in upper trapezius PPT after a 90-second SCS (p<.05). The activity of the upper trapezius and middle deltoid was significantly decreased (p=.014, p=.001), coupled with a decreased muscle activity ratio between the upper and lower trapezius (p<.05). These results indicate that the SCS may effectively deactivate upper trapezius activity, thereby alleviating muscle balance and reducing pain sensitivity.

xylanolytic gut bacterium isolated from Eisenia fetida, Cellulosimicrobium sp. strain HY-13, produced an extracellular glycoside hydrolase capable of efficiently degrading mannose-based substrates such as locust bean gum (LBG), guar gum, mannotetraose, and mannopentaose. The purified mannan-degrading enzyme (ManS, 34,926 Da) from strain HY-13 was found to have an N-terminal amino acid sequence of DEATTDGLHVVDD, which has not yet been identified. Under the optimized reaction conditions of 50℃ and pH 7.0, ManK exhibited extraordinary high specific activities of 7,109 IU/mg and 5,158 IU/mg toward LBG and guar gum, respectively, while the enzyme showed no effect on sugars substituted with p-nitrophenol and various non-mannose carbohydrates. ManK strongly attached to Avicel, lignin, β-cyclodextrin, and poly(3-hydroxybutyrate) granules, but not bound to chitin, chitosan, curdlan, or insoluble oat spelt xylan. The aforementioned characteristics of ManS suggest that it is a unique endo-β -1,4-mannanase with out additional carbohydrolase activities, which differentiates it from other well-known carbohydrolases.

Lentinula edodes is an important cultivated mushroom in China. The development of Lentinula edodes production promotes more studies on it. In our previous work, degenerate PCR and chromosome walking technologies were used to obtain one pheromone receptor gene and one pheromone precursor gene from Lentinula edodes. In this study, four pairs of specific primers were designed according to the whole genome sequencing of the protoplast monokaryon of Lentinula edodes strain 135, to amplify STE3-like pheromone receptor gene and its flanking conserved genes in the protoplast monokaryon strain SUP2 derived from Lentinula edodes strain Suxiang and 33655bp DNA sequence was obtained. By BlastX search, seven putative genes were identified, and three of them are pheromone receptor encoded genes. Furthermore, near to two pheromone receptor genes, four genes encoding proteins with conserved motifs of pheromone precursors were found. This study firstly reveals the molecular organization of the B mating type locus of Lentinula edodes.

[Aims] Lentinula edodes (Berk.) Pegler has many commercial strains both morphologically and physiologically similar to each other. At present, detection of polymorphism in rDNA-IGS region (Babasaki, 2006) and/or RAPD marker (Zhang and Molina, 1995) is generally used for strain typing of L. edodes. However, it is rather time-and-cost consuming. Inter-retrotransposon amplified polymorphism (IRAP)-PCR method mainly used for horticultural crops takes less time and lower in cost in strain typing (Kalendar et al, 1999). In this study, we designed IRAP primers for L. edodes and verified their strain typing efficiency. [Method] Thirty three strains were provided for this study. Either fungal cultures on PDA or fungal tissues of fruit bodies were cut into approximately 4 x 4 x 4 mm. Total DNA of each samples were extracted by DNeasy Plant Mini Kit (QIAGEN). For PCR, IRAP primer set and Pfu-X polymerase (greiner) were performed. Based on LTR (Long Terminal Repeat) sequence in L. edodes, we designed one set of primers amplifying the regions between retrotransposons. Ampricons were electrophored for 50 min at 100 V on 1.7 % agarose gel with GelRed (Biotium) and evaluated under UV irradiation. [Results] The products obtained by IRAP-PCR were determined using mini-gel electrophoresis system. The band patterns of IRAP-PCR products differ among strains except the ones having the same parental cultivar. The detected bands were bright and clear without smearing. The IRAP-PCR products of fungal cultures on PDA and correlating fungal tissues of fruit bodies showed the same band pattern, suggesting that the procedure is highly reproducible. Thus, it is considered that IRAP-PCR with short ranged (ca. 1 kb) electrophoresis is a time-efficient and practical strain typing method of L. edodes.

콩은 식물성 단백질 및 지방의 주요 공급원이고 콩 종실에는 기능성 성분이 많이 함유되어져 있어 소비 가 점차 증가하고 있지만 Lipoxygenase, Kunitz Trypsin inhibitor, 7S α′-subunit 단백질과 같은 성 분들이 존재하는데 이는 품질과 영양가치를 떨어뜨리고 섭취시 알러지를 일으키기도 한다. 콩에서 유전적 으로 이러한 성분이 결핍되어져 있는 유전자형의 선발은 품질이 우수한 콩 육종의 기초단계이다. “개척2 호”와 PI506876의 교배로부터 434개의 F2 종자를 얻어 F2 종자의 일부를 사용하여 SDS-PAGE로 각각 의 종자를 분석한 결과 Lipoxygenase와 Kunitz Trypsin inhibitor 및 7S의 α′-subunit 단백질이 모두 결핍되어져 있는 lx1lx1lx2lx2lx3lx3titicgy1cgy1 유전자형을 가진 종자를 선발하여 F2 식물체로 길러 성 숙 후 F3 종자를 수확하였다. F3 종자로부터 Lipoxygenase, Kunitz Trypsin inhibitor, 7S α′-subunit 단백질이 모두 결핍되어져 있음을 재확인하였으며 선발된 종자는 고품질 콩 품종 육성에 유용하게 활용될 것으로 기대된다.

A new commercial strain "Dan Bi" of Pleurotus eryngii was developed by intra-specific crossing between monokaryotic spores derived from KNR2312 and KNR2596, respectively. The optimum temperature of mycelial growth was 25℃ and that of fruiting body development was 15 -16℃. The primordia formation of "Dan Bi" was one day later than that of control strain KNR2312. The color of pileus and stipe surface was neutral-brown and pure white, respectively. The shape of pileus was dome and has a scale like as cobweb. The yield was 93±4.8g/850cc plastic bottle. Analysis of the genetic characteristics of the new commercial strain "Dan Bi" showed a different profile as that of the control strain KNR2312 when RAPD primer 8001, 8002, 8003, 8004, and 8005 were used. This new variety of King Oyster mushroom is characterized by a few primordia formation after scratching. It would be of much help to mushroom cultivators in a process of culling of P. eryngii.

Pleurotus ostreatus 'Miso' is a mutant strain showing white color in pileus from the known parent strain 'Wonhyeong No. 1'. Shape and several other characters also vary with culture conditions. Mating experiments were performed to understand interstrain mating relationship using monokaryons of the parent and the mutant strains. All monokaryons were grown from single spores isolated from freshly collected fruit bodies. Pairings were performed in 90 mm petri dishes on PDA. They were allowed to grow at 25℃ until two fronts of the advancing mycelia met and developed a conspicuous contact zone. The contact zone and the outer edges of paired colonies on each plate were examined for clamp connections. The crosses were considered positive when clamp connections were at the growing margin of either side of the interacting monokaryons. The parent strain 'Wonhyeong No. 1' and the mutant strain 'Miso' resulted in tetrapolar incompatibility system in intrastrain crosses. Normal compatible matings could be visibly distinguished from incompatible ones by the rapid growth and gross morphology. The resultant dikaryotic colony was also confirmed microscopically by the presence of clamp connections from incompatible pairings. In interstrain crosses, each monokaryotic tester strains of the parent strain was out-crossed to monokaryotic tester strains of the mutant. The four classes of tester strains representing the four incompatibility types were chosen from each mutant and parent strains. As a result of these crosses it was found that both strains share the same A and B incompatibility factors yielding 25% compatibility.

RC교량의 내진성능은 교각에 충분한 연성도를 제공함으로써 확보할 수 있다. 이러한 연성도는 교각의 소성힌지 영역에 적절한 횡방향철근을 배근함으로써 실현할 수 있다. 횡방향철근에 의한 횡구속력은 유효구속력으로 결정되므로 단면형상과 횡방향철근량이 지배적인 요소가 된다. 동일한 횡방향철근량을 제공하더라도 설치간격, 배치형태, 갈고리 상세 등의 차이에 의해 유효구속력에 차이가 있게 된다. 후프띠철근에 의해 횡구속력을 발휘하는 원형단면과는 달리 사각 또는 중공사각단면에서는 유효구속력을 증가시키기 위해 보강띠철근이 함께 사용된다. 이러한 보강띠철근을 어떻게 고려하느냐에 따라 횡구속된 콘크리트의 응력-변형률 관계는 달라지게 된다. 본 연구에서는 실험을 통해 후프띠철근과 함께 보강띠철근을 갖는 정사각단면 콘크리트의 응력-변형률 관계를 파악하였으며 기존의 평가식과 비교를 통해 역학적 특성을 분석하였다.

복합적층구조의 층간분리현상은 탄성좌굴하중을 감소시키며 설계값보다 낮은 수준에서 전체구조물의 파괴를 유발 한다. 따라서 복합적층구조의 층간분리 현상은 매우 중요한 문제이며 많은 이론과 실험적인 연구가 진행되어왔다. 본 연구에서는 3차원 이론을 사용한 효과적인 유한요소법에 기초하여 임베디드된 사각형 층간분리 현상을 갖는 복 합적층판의 탄성좌굴 거동을 분석하였다. 해석을 위해 개발된 3차원 유한요소는 EAS-SOLID8이라고 이름 붙여졌으 며 강화된 대체 변형률 방법을 사용하였다. 임베디드된 사각형 층간분리를 갖는 복합적층판의 탄성좌굴거동 분석을 위해 경계조건, 폭-두께비 변화에 대하여 매개변수 해석을 수행하였다. 본 연구의 그래프와 좌굴모드는 임베디드된 사각형 층간분리를 갖는 복합적층판의 설계에 매우 유용한 자료가 될 것으로 사료된다.

Bacillus thuringiensis 1-3 (Bt 1-3) which was isolated from a Korean soil sample showed high insecticidal activity against Aedes aegypti as well as Plutella xylostella. The isolate was determined to belong to ssp. aizawai (H7) type by an H antiserum agglutination test and produced bipyramidal-shaped crystal proteins with a molecular weight of 130 kDa. PCR analysis with cry gene specific primers showed that Bt 1-3 contained cry1Aa, cry1Ab, cry1C, cry1D and cry2A gene, differing from spp. aizawai (reference strain) which contains cry1Aa, cry1Ab, cry1C and cry1D. We modified the plasmid capture system (PCS) to clone plasmid from Bt 1-3 through in vitro transposition. Fifty-three clones were acquired and their sizes were approximately 10 kb. Based on the sequence analysis, they were classified according to similarities with four known Bt plasmids, pGI3, pBMB175, pGI1 and pGI2, respectively. One of pGI3-like clones, named as pBt1-3, was fully sequenced and its 20 putative open reading frames (ORFs), Rep-protein, double-strand origin of replication (dso), single-strand origin of replication (sso), have been identified. The structure of pBt1-3 showed high similarity with pGI3 which is one of rolling-circle replication (RCR) group VI family.

A new Bacillus thuringiensis isolate 19-22 (Bt 19-22) exhibited high anti-fungal activity against barley powdery mildew (Blumeria graminis f. sp. hordei). The cry gene content of Bt 19-22 comprised cry1Aa, cry1Ab, cry1Ac and cry1D which have high insecticidal activity against lepidopteran larvae. We tried to confer a dipteran insecticidal activity to Bt 19-22 for constructing a recombinant strain which has multiple functions, anti-fungal and dual insecticidal activity. The insecticidal cry11Aa gene of B. thuringiensis was constructed under cry1Ac promoter in an E. coli-B. thuringiensis shuttle vector (pPro11A). The plasmid, pPro11A was introduced into Bt 19-22 isolate by electroporation and four transformants which had different cry gene contents were identified by PCR with cry11Aa and cry1-type specific primers. Among them, a Bt 19-22 transformant (11A/19-22 No. 7) expressed Cry11A protein (approximately 70 kDa) successfully without change of its inherent characteristics such as Cry protein expression and antifungal activity. The insecticidal activity of 11A/19-22 No. 7 was checked against Plutella xylostella and Culex pipiens. These results suggests that the recombinant strain shows dual insecticidal activity against lepidopteran and dipteran larvae as well as antifungal activity.

The silkworm (Bombyx mori), as an industrial insect, possesses a high economic value. Casual discrimination and accumulated genetic information of silkworm varieties are essential ground for the practical utilization and long-term conservation. In this study, nine available microsatellite loci were successfully genotyped from ~50 silkworm strains preserved in Korea. According to genotyping analysis, we obtained 3 ~ 16 alleles per locus, with an average of 7.4, the observed heterozygosity ranging from 0.04 to 0.98, and the polymorphic information content (PIC) ranging from 0.06 to 0.88, revealing that some loci are highly variable. Among 54 strains 13 strains were casually identified by the presence of 17 strain-specific apomorphic alleles. Furthermore, 30 among remaining strains contained strain-specific allele combinations that are also apomorphic to each strain, allowing us to discriminate each of these from other strains by genotyping of multiple loci. These results collectively suggest that the silkworm microsatellite DNA is actually and potentially important molecular marker for the discrimination of the silkworm strains that are preserved as hundreds in Korea, as more loci are genotyped.

The Organophosphorus pesticides are widely used for agricultural and domestic purposes due to their relatively low persistence in the environment. Chlorpyrifos-methyl (CM) is used at a rate of over 14 million pounds per year in US agriculture, ranking it as the second most heavily used pesticide. This study aimed at isolating bacteria from soil and determining their ability to degrade CM and identify the intermediates in culture broth. Bacteria capable of degrading CM was isolated by enrichment culture. Chryseobacterium sp. strain KR200 degraded CM up to 91.58% in 7days. Studies with CM in liquid culture of Chryseobacterium sp. strain KR200 demonstrated that the isolate hydrolyzed CM to 3,5,6-trichloro-2-pyridinol, and utilized this compound for growth and energy. We performed SDS-PAGE and two-dimensional gel electrophoresis and identified proteins whose expression pattern is affected by CM using mass spectrometry. The results revealed various proteins that can be grouped according to their respective cellular function. These results highlight the potential of this bacterium to be used in the clean up of contaminated pesticide waste in the environment.