• Korean Journal of Microbiology
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• v.51 no.1
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• pp.68-74
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• 2015

Based on the genomic analysis of Thermococcus pacificus P-4, we identified a putative GH1 ${\beta}$-glucosidase-encoding gene (Tpa-glu). The gene revealed a 1,464 bp encoding 487 amino acid residues, and the deduced amino acid residues exhibited 77% identity with Pyrococcus furiosus ${\beta}$-glucosidase (accession no. NP_577802). The gene was cloned and expressed in Escherichia coli system. The recombinant protein was purified by metal affinity chromatography and characterized. Tpa-Glu showed optimum activity at pH 7.5 and $75^{\circ}C$, and thermostability with a half life of 6 h at $90^{\circ}C$. Tpa-Glu exhibited hydrolyzing activity against various pNP-glycopyranosides, with kcat/Km values in the order of pNP-${\beta}$-glucopyranoside, pNP-${\beta}$-galactopyranoside, pNP-${\beta}$-mannopyranoside, and pNP-${\beta}$-xylopyranoside. In addition, the enzyme exhibited exo-hydrolyzing activity toward ${\beta}$-1,3-linked polysaccharide (laminarin) and ${\beta}$-1,3- and ${\beta}$-1,4-linked oligosaccharides. This is the first description of an enzyme from hyperthermophilic archaea that displays exo-hydrolyzing activity toward ${\beta}$-1,3-linked polysaccharides and could be applied in combination with ${\beta}$-1,3-endoglucanase for saccharification of laminarin.

• Korean Journal of Microbiology
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• v.44 no.3
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• pp.203-211
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• 2008

Rapid detection-method for Shiga toxin type 1 that was produced from Shiga toxin-producing Escherichia coli (STEC) was developed by two-step ultra-rapid real-time (URRT) PCR. The specific primers were deduced from 80 bp stable region of stx type 1 (stxl) gene among various informations of STEC strains. URRT PCR is a microchip-based real-time PCR using 6 ${\mu}l$ of reaction volume with extremely short denaturation step and annealing/extension step (1 sec, 3 sec, respectively) in each cycle of PCR. Using the stx1-specific URRT PCR, 35 cycled PCR were finished in time of 6 min and 38 see, also measured 7 min and 28 see including melting temperature (Tm) analysis. The detection-limit of stxl-specific URRT-PCR was estimated until 3 colony forming units / PCR with products with stable Tm at $81.42{\pm}0.34^{\circ}C$. In the applications to various STEC strains and contaminated genomic DNAs, stx1-specific URRT-PCR were tested and shown that it would be expected an useful method for the rapid detection of stx1-coded STEC strains.

• Applied Biological Chemistry
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• v.39 no.2
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• pp.112-117
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• 1996

Arginine decarboxylase (ADC) is the first enzyme in one of the two pathways of diamine putrescine biosynthesis in plants. The genes encoding ADC have previously been cloned from Escherichia coli, oat and tomato genome. Two degenerate oligonucleotides (17-mer) corresponding to two conserved regions of ADC were used as primers in polymerase chain reaction of rice (Oryza sativa L.) genomic DNA, and an approximately 1.0 kbp fragment was obtained. This amplified PCR product showed an open reading frame which contains 1,022 bp of nucleotide sequences. This PCR product was cloned into pGEM-originated T vector and the short 500 bp PstI digested fragment was subcloned into pGEM-3zf(+/-) vectors to facilitate sequencing. The nucleotide sequence of this PCR product showed about 74% and 70% identity with the same regions of the oat and tomato ADC cDNA sequences, respectively. The predicted amino acid sequence exhibited 45% and 62% identity with oat and tomato ADC polypeptide fragments, respectively. The sequence similarities of 34%, 47% and 38% were previously reported in oat and E. coli, tomato and oat, and tomato and E. coli ADC amino acids, respectively. Therefore, similarities and identities between rice and oat or tomato are remarkably higher than those others of the previous reports. In the highly conserved regions in both the amino acid sequence and spacing regions among the sequences of these three, rice ADC open reading frame also has the exactly same regions with the striking similarity. RNA blot analysis showed that hnc is expressed as a transcript of approximately 2.5 kbP in the rice seedling leaf tissues.

• Journal of Life Science
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• v.30 no.7
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• pp.614-624
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• 2020

The recent development of next-generation sequencing technology has enabled increased genomic analysis, but very few single nucleotide polymorphism (SNP) markers applicable to sweet persimmon (Diospyros kaki Thunb.) cultivars have been identified. In this study, SNP primers developed from five pollination-constant astringent (PCA) persimmons native to Korea were applied to discriminate between cultivars and verify their usability. The polymerase chain reactions of 19 SNP primers developed by Jung et al. were checked, with 11 primers finally selected. The other eight were very difficult to analyze in the agarose gel electrophoresis and QIAxcel Advanced System used in this experiment and were therefore excluded. The 11 SNP primers were applied through first and second verification to 76 cultivars and collection lines including 20 pollination-variant non-astringent (PVNA), 30 pollination-constant non-astringent (PCNA), 20 PCA, and six pollination-variant astringent (PVA). Of these, 38 were indistinguishable (eight PVNA, 18 PCNA, nine PCA, and three PVA). However, the results of applying the 11 SNP primers to new sweet persimmon cultivars, namely Gamnuri, Dannuri, Hongchoo, Jamisi, and Migamjosaeng, showed that they have the potential to be used as a unique marker for simultaneously determining between them.

• Clinical and Experimental Reproductive Medicine
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• v.29 no.4
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• pp.303-310
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• 2002

Objective s: To estimate the frequency of Y chromosome microdeletions in the Korean population of infertile men and to evaluate the relationship between microdeletion on the Y chromosome and clinical phenotypes of infertile men with idiopathic azoospermia and oligozoospermia. Materials and Methods: Genomic DNA was extracted from blood samples collected from 330 infertile men attending the Infertility Clinic at Samsung Cheil Hospital, Korea. Six sequence tagged sites (STSs) spanning the azoospermia factor (AZF) regions of the Y chromosome were amplified by polymerase chain reactions (PCRs). Results: Microdeletions on Y chromosome were detected in 35 (10.6%) of the 330 infertile men. Most of the microdeletions (91.4%) involved AZFb or AZFc. The high incidence of microdeletions were found in AZFc region (57.1%), but the low in AZFa (8.6%) and AZFb (5.7%). Larger microdeletions involving two or three AZF regions were detected in 28.6% of cases. All patients (6 patients) with deletion of AZFa region showed no germ cell phenotypes, Sertoli cell only syndrome or Leydig cell hyperplasia in histopathologic examinations. Conclusion: Microdeletions on the Y chromosome, especially, at AZFc/DAZ regions may be the major cause of azoospermia and severe oligozoospermia. We suggest that idiopathic infertile men have genetic counselling and microdeletion analysis on the Y chromosome before IVF-ET and ART program.

• Journal of Marine Bioscience and Biotechnology
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• v.1 no.3
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• pp.186-192
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• 2006

Short interspersed repetitive elements (SINEs) are dispersed throughout eukaryotic genomes. These SINEs have been shown to be excellent phylogenetic markers for the closed related species. In this report, we isolated two novel families of SINEs from the mud loach. The two SINE families, mlSINE-L and mlSINE-S, have genomic lengths of about 410bp and 270bp, respectively. 5' and 3' ends of the SINE families are well conserved and highly homologous to each of corresponding ends of RSg-1 and SmaI SINEs. Phylogenetic analysis shows that mlSINEs are unique to the mud loach. A dot blot hybridization experiment shows that mlSINE-L has an estimated copy number of $1{\times}10^3$ per $2{\times}10^9bp$ (2.8 pg) and is more frequently distributed at nuclear matrix attachment regions (MARs) than loop DNAs. The result suggests that mlSINEs may preferentially integrate in or near MARs.

• Molecules and Cells
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• v.38 no.11
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• pp.950-958
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• 2015

BCS1L gene encodes mitochondrial protein and is a member of conserved AAA protein family. This gene is involved in the incorporation of Rieske FeS and Qcr10p into complex III of respiratory chain. In our previous study, AluYRa2-derived alternative transcript in rhesus monkey genome was identified. However, this transcript has not been reported in human genome. In present study, we conducted evolutionary analysis of AluYRa2-exonized transcript with various primate genomic DNAs and cDNAs from humans, rhesus monkeys, and crabeating monkeys. Remarkably, our results show that AluYRa2 element has only been integrated into genomes of Macaca species. This Macaca lineage-specific integration of AluYRa2 element led to exonization event in the first intron region of BCS1L gene by producing a conserved 3' splice site. Intriguingly, in rhesus and crabeating monkeys, more diverse transcript variants by alternative splicing (AS) events, including exon skipping and different 5' splice sites from humans, were identified. Alignment of amino acid sequences revealed that AluYRa2-exonized transcript has short N-terminal peptides. Therefore, AS events play a major role in the generation of various transcripts and proteins during primate evolution. In particular, lineage-specific integration of Alu elements and species-specific Alu-derived exonization events could be important sources of gene diversification in primates.

• Microbiology and Biotechnology Letters
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• v.38 no.4
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• pp.373-378
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• 2010

Studies on standardization and quality upgrade of nuruk which is a basic component in brewing are required to increase the quality level of Korean traditional rice wines and to develop the technology for practical use of it. It is important to isolate best strains, to improve the properties and effectively preserve them for brewing industry. In this study, 16 commercial nuruk samples were obtained from the commercial markets located in Chungcheong areas in Korea. 174 fungal strains were isolated from the samples on DG18 medium using a dilution plating method and then screened for enzyme activity and acid production. The active strains were identified based on the morphological characteristics and ITS sequence analysis. Out of 174 strains, 12 strains showed high amylase activity. Especially, Rhizopus sp. CN084, CN174, Aspergillus sp. CN161 and Mycocladus sp. CN042 showed high saccharogenic power and dextrinogenic enzyme activity on cooked wheat bran medium. On the other hand, Aspergillus sp. CN010, CN161, Rhizopus sp. CN105, CN168 and Rhizomucor sp. CN088 produced high acid production on the same medium. Our results showed that the active strains may be used as microbial sources for nuruk starter with good quality in brewing.

• Fisheries and Aquatic Sciences
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• v.8 no.3
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• pp.151-160
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• 2005

Genomic DNA samples isolated from Fenneropenaeus chinensis (fleshy prawn; FP) and Palaemon gravieri (Chinese ditch prawn; CDP) collected in the West Sea, off the Korean Peninsula, at Buan, were PCR-amplified repeatedly. The sizes of the DNA fragments generated by seven different primers varied from 50 bp to 1,600 bp. We identified 358 fragments for the FP species and 301 fragments for the CDP species. There were 18 polymorphic fragments (5.03$\%$) for the FP species and 12 (3.99$\%$) for the CDP species. In total, 66 common fragments (average of 9.4 fragments per primer) were observed for the FP species and 44 fragments (average of 6.3 fragments per primer) were observed for the CDP species. The numbers of specific fragments seen for the FP species and CDP species were 38 and 47, respectively. The complexity of the banding patterns varied dramatically between the primers and the two species. In the FP species, a specific fragment of approximately 1,200 bp generated by primer OPB-04 exhibited inter-individual-specific characteristics that were indicative of DNA polymorphisms. Moreover, in the CDP species, a major fragment of approximately 550 bp generated by primer OPB-20 was found to be specific for the CDP. The average bandsharing value between the two prawn species was 0.421$\pm$0.006, and ranged from 0.230 to 0.611. The dendrogram obtained using the data from the seven primers indicated seven genetic clusters: cluster 1, FLESHY 01, 02, 03, and 04; cluster 2, FLESHY 05, 06, and 07; cluster 3, FLESHY 08, 09, 10, and 11; cluster 4, DITCH 13, 14, 16, and 18; cluster 5, DITCH 12, 15, and 17; cluster 6, DITCH 19, 20, and 21; and cluster 7, DITCH 22. The genetic distance between the two prawn species ranged from 0.071 to 0.642. Thus, RAPD-PCR analysis revealed a significant genetic distance between the two prawn species. Using various arbitrary primers, RAPD-PCR may be applied to identify specific/polymorphic markers that are particular to a species and geographic population, and to define genetic diversity, polymorphisms, and similarities among shrimp species.

• Korean Journal of Plant Taxonomy
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• v.41 no.4
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• pp.299-314
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• 2011

The choice of molecular markers is the first step when selecting experimental plans in the field of population genetics. The popular molecular markers in population genetic studies are mainly allozyme, RAPD, RFLP, AFLP, microsatellite, SNP and ISSR. Among these, microsatellites are frequently found in nuclear, chloroplast and mitochondrial genome, showing a high level of polymorphism and nuclear microsatellites are codominant. Thus, it is a favorable molecular marker for population structure analyses and genetic diversity studies. Microsatellites are composed of tandem repeated 1~6 base pair nucleotide motifs and can be easily amplified by PCR reactions using locus specific primers. Because microsatellites have low cross-species transferability, however, they are only applicable between phylogenetically close species. In wild plants, the lack of genomic information and the high development cost of the microsatellite obstruct the wider use of microsatellites in plant population genetics research. In this review, we introduce the basis for microsatellite markers, the development process, and analytical methods as well as evolutionary models and their applications. In addition, possible genotyping errors which lead to erroneous conclusions are discussed.