• Journal of Microbiology and Biotechnology
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• 제24권9호
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• pp.1196-1206
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• 2014

A metagenomic fosmid library was constructed using genomic DNA isolated from the gut microflora of Hermetia illucens, a black soldier fly. A cellulase-positive clone, with the CS10 gene, was identified by extensive Congo-red overlay screenings for cellulase activity from the fosmid library of 92,000 clones. The CS10 gene was composed of a 996 bp DNA sequence encoding the mature protein of 331 amino acids. The deduced amino acids of CS10 showed 72% sequence identity with the glycosyl hydrolase family 5 gene of Dysgonomonas mossii, displaying no significant sequence homology to already known cellulases. The purified CS10 protein presented a single band of cellulase activity with a molecular mass of approximately 40 kDa on the SDS-PAGE gel and zymogram. The purified CS10 protein exhibited optimal activity at $50^{\circ}C$ and pH 7.0, and the thermostability and pH stability of CS10 were preserved at the ranges of $20{\sim}50^{\circ}C$ and pH 4.0~10.0. CS10 exhibited little loss of cellulase activity against various chemical reagents such as 10% polar organic solvents, 1% non-ionic detergents, and 0.5 M denaturing agents. Moreover, the substrate specificity and the product patterns by thin-layer chromatography suggested that CS10 is an endo-${\beta}$-1,4-glucanase. From these biochemical properties of CS10, it is expected that the enzyme has the potential for application in industrial processes.

• Asian-Australasian Journal of Animal Sciences
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• 제20권5호
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• pp.768-774
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• 2007

To study the effect of dietary docosahexaenoic acid (DHA) enrichment on the expression of hepatic genes in pigs, weaned, crossbred pigs (30 d old) were fed diets supplemented with either 2% tallow or DHA oil for 18 d. Hepatic mRNA was extracted. Suppression subtractive hybridization was used to explore the hepatic genes that were specifically regulated by dietary DHA enrichment. After subtraction, we observed 288 cDNA fragments differentially expressed in livers from pigs fed either 2% DHA oil or 2% tallow for 18 d. After differential screening, 7 genes were found to be differentially expressed. Serum amyloid A protein 2 (SAA2) was further investigated because of its role in lipid metabolism. Northern analysis indicated that hepatic SAA2 was upregulated by dietary DHA enrichment (p<0.05). In a second experiment, feeding 10% DHA oil for 2d significantly increased the expression of SAA2 (compared to the 10% tallow group; p<0.05). The porcine SAA2 full length cDNA sequence was cloned and the sequence was compared to the human and mouse sequences. The homology of the SAA2 amino acid sequence between pig and human was 73% and between pig and mouse was 62%. There was a considerable difference in SAA2 sequences among these species. Of particular note was a deletion of 8 amino acids, in the pig compared to the human. This fragment is a specific characteristic for the SAA subtype that involved in acute inflammation reaction. Similar to human and mouse, porcine SAA2 was highly expressed in the liver of pigs. It was not detectable in the skeletal muscle, heart muscle, spleen, kidney, lung, and adipose tissue. These data suggest that SAA2 may be involved in mediation of the function of dietary DHA in the liver of the pig, however, the mechanism is not yet clear.

• Journal of Microbiology and Biotechnology
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• 제19권2호
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• pp.113-120
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• 2009

Twenty-seven fenitrothion-degrading bacteria were isolated from different soils, and their genetic and phenotypic characteristics were investigated. Analysis of the 16S rDNA sequence showed that the isolates were related to members of the genera Burkholderia, Pseudomonas, Sphingomonas, Cupriavidus, Corynebacterium, and Arthrobacter. Among the 27 isolates, 12 different chromosomal DNA fingerprinting patterns were obtained by polymerase chain reaction(PCR) amplification of repetitive extra genic palindromic(REP) sequences. The isolates were able to utilize fenitrothion as a sole source of carbon and energy, producing 3-methyl-4-nitrophenol as the intermediate metabolite during the complete degradation of fenitrothion. Twenty-two of 27 isolates were able to degrade parathion, methyl-parathion, and p-nitrophenol but only strain BS2 could degrade EPN(O-ethyl-O-p-nitrophenyl phenylphosphorothioate) as a sole source of carbon and energy for growth. Eighteen of the 27 isolates had plasmids. When analyzed with PCR amplification and dot-blotting hybridization using various specific primers targeted to the organophosphorus pesticide hydrolase genes of the previously reported isolates, none of the isolates showed positive signals, suggesting that the corresponding genes of our isolates had no significant sequence homology with those of the previously isolated organophosphate pesticide-degrading bacteria.

• 한국미생물·생명공학회지
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• 제36권2호
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• pp.165-172
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• 2008

본 연구에서는 분리균주 LH2를 이용하여 각각의 중금속 (Cr, Cu, Ni, Pb, Zn)에 대한 최적성장조건을 도출하였으며, pH, 온도 및 중금속 농도 변화에 따른 중금속 제거효율을 관찰하였다. 분리균주는 18S rDNA 분석에 의거하여 종 유사성이 100%로서 Exophiala sp.로 동정되었다. 분리균주의 경우 $25^{\circ}C$, pH 7인 진탕배양 조건에서 가장 높은 제거효율을 보였다. 첨가되는 중금속의 농도가 200 ppm이하일 경우 중 금속의 비제거속도를 구한 결과 pH 7인 조건에서 중금속의 종류와 무관하게 10 ppm에서 200 ppm으로 중금속의 농도가 증가함에 따라 0.01에서 4.43 mg-metal $L^{-1}{\cdot}d^{-1}{\cdot}mg^{-1}{\cdot}DCW^{-1}$으로 증가하였다. pH 7로 설정된 배양액에 약 200ppm의 중금속이 첨가될 경우 중금속의 제거효율은 Cr, Cu, Ni, Pb, 및 Zn 이 각각 99.28%, 97.67%, 91.94%, 99.77%, 99.61%로 관찰되었다.

• Parasites, Hosts and Diseases
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• 제54권6호
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• pp.751-758
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• 2016

This study aimed at constructing a draft genome of the adult female worm Toxocara canis using next-generation sequencing (NGS) and de novo assembly, as well as to find new genes after annotation using functional genomics tools. Using an NGS machine, we produced DNA read data of T. canis. The de novo assembly of the read data was performed using SOAPdenovo. RNA read data were assembled using Trinity. Structural annotation, homology search, functional annotation, classification of protein domains, and KEGG pathway analysis were carried out. Besides them, recently developed tools such as MAKER, PASA, Evidence Modeler, and Blast2GO were used. The scaffold DNA was obtained, the N50 was 108,950 bp, and the overall length was 341,776,187 bp. The N50 of the transcriptome was 940 bp, and its length was 53,046,952 bp. The GC content of the entire genome was 39.3%. The total number of genes was 20,178, and the total number of protein sequences was 22,358. Of the 22,358 protein sequences, 4,992 were newly observed in T. canis. Following proteins previously unknown were found: E3 ubiquitin-protein ligase cbl-b and antigen T-cell receptor, zeta chain for T-cell and B-cell regulation; endoprotease bli-4 for cuticle metabolism; mucin 12Ea and polymorphic mucin variant C6/1/40r2.1 for mucin production; tropomodulin-family protein and ryanodine receptor calcium release channels for muscle movement. We were able to find new hypothetical polypeptides sequences unique to T. canis, and the findings of this study are capable of serving as a basis for extending our biological understanding of T. canis.

• BMB Reports
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• 제38권4호
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• pp.420-431
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• 2005

The differentially virulent race T1 of common bunt (Tilletia tritici) was used to inoculate the wheat lines Neepawa (compatible) and its sib BW553 (incompatible) that are nearly isogenic for the Bt-10 resistance gene. Inoculated crown tissues were used to construct a suppression subtractive hybridization (SSH) cDNA library. Of the 1920 clones arrayed from the SSH cDNA library, approximately 10% were differentially regulated. A total of 168 differentially up-regulated and 25 down-regulated genes were identified and sequenced; 71% sequences had significant homology to genes of known function, of which 59% appeared to have roles in cellular metabolism and development, 24% in abiotic/biotic stress responses, 3% involved in transcription and signal transduction responses. Two putative resistance genes and a transcription factor were identified among the up regulated sequences. The expression of several candidate genes including a lipase, two non-specific lipid transfer proteins (ns-LTPs), and several wheat pathogenesis-related (PR)-proteins, was evaluated following 4 to 32 days post-inoculation in compatible and incompatible interactions. Results confirmed the higher overall expression of these genes in resistant BW553 compared to susceptible Neepawa, and the differential up-regulation of wheat lipase, chitinase and PR-1 proteins in the expression of the incompatible interaction.

• Asian-Australasian Journal of Animal Sciences
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• 제25권6호
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• pp.758-763
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• 2012

As a member of a subclass of immunophilins, it is controversial that FKBP38 acts an upstream regulator of mTOR signaling pathway, which control the process of cell-growth, proliferation and differentiation. In order to explore the relationship between FKBP38 and mTOR in the Cashmere goat (Capra hircus) cells, a full-length cDNA was cloned (GenBank accession number JF714970) and expression pattern was analyzed. The cloned FKBP38 gene is 1,248 bp in length, containing an open reading frame (ORF) from nucleotide 13 to 1,248 which encodes 411 amino acids, and 12 nucleotides in front of the initiation codon. The full cDNA sequence shares 98% identity with cattle, 94% with horse and 90% with human. The putative amino acid sequence shows the higher homology which is 98%, 97% and 94%, correspondingly. The bioinformatics analysis showed that FKBP38 contained a FKBP_C domain, two TPR domains and a TM domain. Psite analysis suggested that the ORF encoding protein contained a leucine-zipper pattern and a Prenyl group binding site (CAAX box). Tissue-specific expression analysis was performed by semi-quantitative RT-PCR and showed that the FKBP38 expression was detected in all the tested tissues and the highest level of mRNA accumulation was detected in testis, suggesting that FKBP38 plays an important role in goat cells.

• Journal of Microbiology and Biotechnology
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• 제13권2호
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• pp.207-216
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• 2003

A number of different sources, such as composts, leachates, and pig feces samples, were collected from different pig farms in Korea, and several microorganisms were screened for their ability to deodorize the malodorous gases. Consequently, a novel malodorous gases-deodorizing bacterial strain, KJ-108. was isolated, because it was highly abundant in nitrate-supplemented minimal medium ($MM-NO_3^-$) under anaerobic culture conditions. Airtight crimp-sealed serum bottles containing $MM-NO_3^-$ , medium were inoculated with KJ-108. Nitrate concentration was decreased rapidly after 20 h of incubation, and incubation was carried out until nitrite production reached almost zero. Taxonomic identification, including 16S rDNA base sequencing and phylogenetic analysis, indicated that the isolate had $100\%$ homology in its 165 rDNA base sequence with Lactobacillus pentosus. Among the volatile fatty acids, acetic acid contained in large amounts in fresh piggery slurry was decreased by about $40\%$ after 50 h incubation with strain KJ-108. n-Butyric acid, n-valeric acid, and isovaleric acid were gradually decreased, and isobutyric acid and capronic acid were dramatically eliminated at theinitial period with the treatment. Moreover, NH, removal efficiency reached a maximum of $98.5\%$ after 50 h of incubation, but the concentration of $H_2S$ was not changed.

• 한국미생물·생명공학회지
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• 제32권2호
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• pp.172-178
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• 2004

본 연구는 배추 무사마귀병의 생물학적 방제를 위해 수행되었다. 배추 무사마귀병균 Plasmodiophora sp외 길항미생물은 낙엽부식토양에서 분리한 350여 종의 토양미생물 중에서 가장 길항력이 높은 방선균 AC-3균주를 이용하였고, AC-3 균주는 16S rDNA 염기서열분석방법으로 Streptomyces sp.로 동정되었다. Streptomyces sp. AC-3은 배양액 $m\ell$당 9.3 units의 chitinase를 생산하였다. 그 결과 배추 무사마귀병균 Plasmodiophora sp.의 배양액에 Streptomyces sp. AC-3을 접종하고 배양하면 Plasmodiophora sp.의 균체가 팽윤되거나 세포벽이 용해된 모습이 관찰되었다. Streptomyces sp. AC-3은 전딘도 배추 무사마귀병이 만연했던 포장에서 재배 시험하기 위해 제형화시키고 유기질비료에 첨가하여 이용하였다. 미생물제제를 첨가하지 많고 제조한 유기질비료(H-1)(대조구)와 미생물제제 첨가하여 제조한 유기질비료(H-2)(처리구)에 의한 포장시험에서 배추의 생육은 재배 40일 까지는 거의 차이를 나타나지 않았으나 재배 60일에는 다소 차이를 나타내었다. 배추의 병 발생율을 조사한 결과 무사마귀병은 H-1 비료 처리구에서는 36.17% 병이 발생되었으나 H-2 비료 처리구에서는 18.28% 발병되어 약 50%의 방제효과를 나타내었으며, 균핵병, 잎마름병, 그루썩음병 등에서도 길항효과를 나타내어 생물학적 방제가 가능한 균주로 확인되었다.

• Asian-Australasian Journal of Animal Sciences
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• 제28권6호
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• pp.888-895
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• 2015

Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and subsequent sub-cloning and sequencing were used in this study to analyze the molecular phylogenetic diversity and spatial distribution of bacterial communities in different spatial locations during the cooling stage of composted swine manure. Total microbial DNA was extracted, and bacterial near full-length 16S rRNA genes were subsequently amplified, cloned, RFLP-screened, and sequenced. A total of 420 positive clones were classified by RFLP and near-full-length 16S rDNA sequences. Approximately 48 operational taxonomic units (OTUs) were found among 139 positive clones from the superstratum sample; 26 among 149 were from the middle-level sample and 35 among 132 were from the substrate sample. Thermobifida fusca was common in the superstratum layer of the pile. Some Bacillus spp. were remarkable in the middle-level layer, and Clostridium sp. was dominant in the substrate layer. Among 109 OTUs, 99 displayed homology with those in the GenBank database. Ten OTUs were not closely related to any known species. The superstratum sample had the highest microbial diversity, and different and distinct bacterial communities were detected in the three different layers. This study demonstrated the spatial characteristics of the microbial community distribution in the cooling stage of swine manure compost.