• Korean Journal of Veterinary Research
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• v.45 no.1
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• pp.45-53
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• 2005

GroE that is a heat shock protein composed of GroEL and GroES is known as an immunodominant target of both the humoral and cellular immune responses in bovine brucellosis. This study was carried out to characterize groE gene encoding heat shock proteins of B. abortus isolated in Korea and to evaluate the immunogenicity of the GroE protein expressed in E. coli system. In PCR the specific signals with the size of 2,077 bp were detected in five strains isolated from the mammary lymphnodes of the dairy cattle that were serologically positive and the reference strains. In comparison of the sequences of nucleotides and amino acids among the strains, GroES showed 100% identity in both sequences. GroEL was evaluated 99.0~99.9% in nucleotides and 98.0~100% homology in amino acids. The groE gene including groES and groEL was inserted into pET29a vector and constructed pET29a-GroE recombinant plasmids. The inserted groE was confirmed by digestion with Nco1 and EcoR1 endonucleases and nucleotide sequencing. E. coli BL (DE3) was transformed with pET29a-GroE, named as E. coli BL (DE3)/pET29a-GroE. In SDS-PAGE, it was evident that the recombinant plasmid effectively expressed the polypeptides for GroES (10 kDa) and GroEL (60 kDa) in 0.5, 1 and 2 hours after IPTG induction. The immuno-reactivity of the expressed proteins were proved in mouse inoculation and Western blot analysis.

• Korean Journal of Ichthyology
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• v.23 no.2
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• pp.95-105
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• 2011

To explain the origin of the Korean mandarin fish (Siniperca scherzeri), phylogenetic relationships and DNA polymorphism among Siniperca fishes have been investigated based on mitochondrial cytochrome b DNA sequences. As a result, S. roulei were firstly differentiated early in the evolution of Siniperca fishes and the other six species (S. schezeri, S. undulata, S. fortis, S. obscura, S. knerii and S. chuatsi) were evolved slightly later. However, the order of species differentiation among six species was not clear because the nodes of their phylogeny were poorly resolved. The constructed molecular phylogeny revealed three genetically distinct groups of local populations of S. scherzeri. The first group (group 1) is the local populations of Korean peninsula and northern China including Lioaning and Henan. The second one (group 2) is the local populations of Anhui, Fujian and Guangxi. The third one (group 3) is the local population of Zhejiang. The number of nucleotide differences in base pairs were 31~43 between group 1 and 2; 37~44 between group 2 and 3; 27~29 between group 1 and 3; and 1~5 within group 1. Thus, the Korean mandarin fish was likely to be originated from the northern China local population which was isolated from the middle or southern China local populations during the Cenozoic Pliocene. Low level of sequence divergence between Korean mandarin fish populations and northern China population indicated a recent expansion of distribution ranges from northern China to Korean peninsula.

• The Korean Journal of Malacology
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• v.26 no.2
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• pp.185-188
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• 2010

The Web-based the genus Haliotis SNP database was constructed on the basis of Intel Server Platform ZSS130 dual Xeon 3.2 GHz cpu and Linux-based (Cent OS) operating system. Haliotis related sequences (2,830 nucleotide sequences, 9,102 EST sequences) were downloaded through NCBI taxonomy browser. In order to eliminate vector sequences, we conducted vector masking step using cross match software with vector sequence database. In addition, poly-A tails were removed using Trimmest software from EMBOSS package. The processed sequences were clustered and assembled by TGICL package (TIGR tools) equipped with CAP3 software. A web-based interface (Haliotis SNP Database, http://www.haliotis.or.kr) was developed to enable optimal use of the clustered assemblies. The Clustering Res. menu shows the contig sequences from the clustering, the alignment results and sequences from each cluster. And also we can compare any sequences with Haliotis related sequences in BLAST menu. The search menu is equipped with its own search engine so that it is possible to search all of the information in the database using the name of a gene, accession number and/or species name. Taken together, the Web-based SNP database for Haliotis will be valuable to develop SNPs of Haliotis in the future.

• Journal of Life Science
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• v.13 no.3
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• pp.291-297
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• 2003

A human endogenous retroviral family (HERV-W) has recently been described that is related to multiple sclerosis-associated retrovirus (MSRV) sequences that have been identified in particles recovered from monocyte cultures from patients with multiple sclerosis. Two pol fragments (HWP-FB10 and HWP-FBl2) of HERV-W family were identified and analysed by the PCR approach with cDNA library of human fetal brain. They showed 89 percent nucleotide sequence similarity with that of the HERV-W (accession no. AF009668). Deletion/insertion or point mutation in the coding region of the pol fragments from human fetal brain resulted in amino acid frameshift that induced a mutated protein. Phylogenetic analysis of the HERV-W family from GenBank database indicates that the HWP-FB10 is very closely related to the AC000064 derived from human chromosome 7q21-q22. Further studies on the genetic relationship with neighbouring genes and functional role of these new HERV-W pol sequences are indicated.

• Journal of Life Science
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• v.27 no.12
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• pp.1415-1420
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• 2017

The metallothionein (MT) gene (IbMT3) was selected from an EST library of suspension-cultured sweet potato cells. The MT gene, which is one of abundant ESTs in the library, is involved in stress regulation of cells and tissues. A full-length IbMT3 cDNA was obtained and analysis of its nucleotide sequence revealed that IbMT3 encoded a type 3 MT protein, based on its structural characteristics. The function of type 3 MT in plants is not yet known. Northern blot analysis showed stronger expression of IbMT3 in suspension-cultured cells than in sweet potato plant leaves. Since cell culture is known to impose a state of oxidative stress on cells, sweet potato plants were subjected to oxidative stress to investigate the transcriptional regulation of IbMT3. When the herbicide methyl viologen (MV) was administered for 6, 12, and 24 hr, IbMT3 transcription rapidly increased at 6 hr and then decreased. A cold treatment at $15^{\circ}C$ for 24 and 48 hr resulted in a gradual increase in IbMT3 expression. These findings indicate that IbMT3 expression is regulated in response to environmental and oxidative stress. IbMT3 isoform is expected to have antioxidant effects in sweet potato plants and may play an important role in cellular adaptation to oxidative stress.

• Korean Journal of Microbiology
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• v.32 no.4
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• pp.245-251
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• 1994

Ten strains of 7 species from the genus Ganoderma, G. lucidum ATCC 64251, FP-103561-T, and ES70701, G. applanatum ATCC 44053 and FP-57035-T. G. lobatum ATCC 42985, G. resinaceum ATCC 52416, G. subamboinense var. laevisporum ATCC 52420, G. meredithae ATCC 64492, and G. microsporum ATCC 76024, were studied to discuss their phylogenetic relationships by utilizing restriction fragment length polymorphisms (RFLPs) of mitochondrial DNAs (mtDNAs). Six restriction enzymes, BamHI, BglII, EcoRI, HindIII, PvuII, and XbaI which digested mtDNAs into adequate numbers of restriction fragments for cluster analysis, were used in this study. Restriction profiles of strains for each restriction enzyme were treated as analysis characters to calculate similarity coefficients, which were converted into nucleotide sequence divergence values whose mean values were then arranged in a matrix table. This table was utilized for a phylogenetic analysis using the Neighborjoining method of the PHYLIP package to construct phylogenetic tree. Three strains of G. lucidum and two strains of G. applanatum exhibited different lineages each but one of G. applanatum strains showed a close relationship with G. lobatum, which reflected the species complexity of these species whose strains were phenotypically indistinguishable but genetically distinct. The present results suggest that the natural classification of Ganoderma needs to be considered from the viewpoints of molecular biology-based systematics as well as morphological classifications and cultural identifications for better phylogenetic conclusions.

• Korean Journal of Microbiology
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• v.44 no.4
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• pp.339-345
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• 2008

This study was to detect MBL (metallo-${\beta}$-lactamase) among glucose non-fermenting Gram-negative bacilli isolated from clinical specimen and to search antimicrobial combination therapy against MBL producing Pseudomonas aeruginosa. Among fifty one isolates of Gram-negative bacilli with reduced imipenem susceptibility ($MIC{\ge}8{\mu}g/ml$), nine isolates have shown positive results in MBL detection test. They were seven Achromobacter xylosoxidans subsp. xylosoxidans and two P. aeruginosa. The results from EDTA-DDST coin-cided with those of PCR and nucleotide sequence analysis which showed the presence of $bla_{VIM-2}$. The combination of aztreonam (AZT) and piperacillin-tazobactarn (TZP) or AZT and amikacin (AN) screened by one disk synergy test showed no synergistic effect. Triple antibiotic combination therapy with AZT, TZP and AN, however, was shown to be effective and the most synergistic after 8 hrs of exposure. This result strongly suggest that the triple combination therapy of AZT, TZP, and AN could be useful for the treatment of infection caused by MBL producing Gram-negative bacilli.

• KSBB Journal
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• v.16 no.2
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• pp.188-199
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• 2001

A gene encoding the dextransucrase(dsCB) that synthesizes mostly $\alpha-(1\rightarrow6)$ linked dextran with low amount(10%) of $\alpha-(1\rightarrow3)$ branching was cloned and sequenced from Leuconostoc mesenteroides B-742CB. The 6.1 kbp DNA fragment carrying dsCB showed one open reading frame(ORF) composed of 4,536bp. The deduced amino acid sequence shows that it begins from the start codon(ATG) at position 698 of the cloned DNA fragment and extends to the termination condon(TAA) at position 5,223. The enzyme is consisted of 1,508 amino acids and has an calculated molecular mass of 168.6kDa. This calculated Mw was in good agreement with an activity band of 170kDa on non-denaturing SDS-PAGE. A recombinant E. coli DH5 $alpha$ harboring pDSCB produced extracellular dextransucrase in 2% sucrose medium, and synthesized both soluble and insoluble dextran. To compare the properties of enzyme with B-742CB dextransucrase, the acceptor reaction, hydrolysis of dextran and methylation were performed. The expressed enzyme showed the same properties as B-742CB dextransucrease, but its ability to synthesize $\alpha-(1\rightarrow3)$ branching was lower than that of B-742CB dextransucrase. In order to identify the critical amino acid residues known as conserved regions related to catalytic activity, Asp-492 was replaced with Asn. D492N resulted in a 1.6 fold decrease in specific activity.

• Journal of Microbiology and Biotechnology
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• v.27 no.9
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• pp.1701-1710
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• 2017

Classical swine fever virus (CSFV) is the etiologic agent of classical swine fever, a highly contagious disease that causes significant economic losses to the swine industry. The lapinized C-strain, a widely used vaccine strain against CSFV, has low growth efficiency in cell culture, which limits the productivity in the vaccine industry. In this study, a recombinant virus derived from C-strain was constructed and subjected to continuous passaging in PK-15 cells with the goal of acquiring a high progeny virus yield. A cell-adapted virus variant, RecCpp80, had nearly 1,000-fold higher titer than its parent C-strain but lost the ability to induce fever in rabbits. Sequence analysis of cell-adapted RecC variants indicated that at least six nucleotide changes were fixed in RecCpp80. Further adaption of RecCpp80 variant in swine testicle cells led to a higher virus yield without additional mutations. Introduction of each of these residues into the wild-type RecC backbone showed that one mutation, M979R (T3310G), located in the C-terminal region of E2 might be closely related to the cell-adapted phenotype. Rabbit inoculation revealed that $RecCpp40_{+10}$ failed to induce fever in rabbits, whereas $RecCpp80_{+10}$ caused a fever response similar to the commercial C-strain vaccine. In conclusion, the C-strain can be adapted to cell culture by introducing specific mutations in its E2 protein. The mutations in RecCpp80 that led to the loss of fever response in rabbits require further investigation. Continuous passaging of the C-strain-based recombinant viruses in PK-15 cells could enhance its in vitro adaption. The non-synonymous mutations at 3310 and 3531 might play major roles in the enhanced capacity of general virus reproduction. Such findings may help design a modified C-strain for improved productivity of commercial vaccines at reduced production cost.

• Mycobiology
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• v.45 no.4
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• pp.379-384
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• 2017

In mating of Lentinula edodes, dikaryotic strains generated from certain monokaryotic strains such as the B2 used in this study tend to show better quality of fruiting bodies regardless of the mated monokaryotic strains. Unlike B2, dikaryotic strains generated from B16 generally show low yields, with deformed or underdeveloped fruiting bodies. This indicates that the two nuclei in the cytoplasm do not contribute equally to the physiology of dikaryotic L. edodes, suggesting an expression bias in the allelic genes of the two nuclei. To understand the role of each nucleus in dikaryotic strains, we investigated single nucleotide polymorphisms (SNPs) in laccase genes of monokaryotic strains to reveal nuclear origin of the expressed mRNAs in dikaryotic strain. We performed reverse transcription PCR (RT-PCR) analysis using total RNAs extracted from dikaryotic strains (A5B2, A18B2, and A2B16) as well as from compatible monokaryotic strains (A5, A18, and B2 for A5B2 and A18B2; A2 and B16 for A2B16). RT-PCR results revealed that Lcc1, Lcc2, Lcc4, Lcc7, and Lcc10 were the mainly expressed laccase genes in the L. edodes genome. To determine the nuclear origin of these laccase genes, the genomic DNA sequences in monokaryotic strains were analyzed, thereby revealing five SNPs in Lcc4 and two in Lcc7. Subsequent sequence analysis of laccase mRNAs expressed in dikaryotic strains revealed that these were almost exclusively expressed from B2-originated nuclei in A5B2 and A18B2 whereas B16 nucleus did not contribute to laccase expression in A2B16 strain. This suggests that B2 nucleus dominates the expression of allelic genes, thereby governing the physiology of dikaryons.