• The Plant Pathology Journal
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• v.19 no.1
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• pp.24-29
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• 2003

SS2-2, a hypernodulating soybean mutant was isolated by EMS mutagenesis from Sinpaldalkong 2. This auto-regulation mutant showed greater number of nodules and smaller plant size than its wild type Sinpaldalkong 2. SSR markers were used to identify DNA variation at SSR loci from different soybean LG. The only SSR marker that detected a length polymorphism between SS2-2 and its wild type ancestor was Satt294 on LG C1 instead of LG H, locating a hypernodulating gene. Sequencing data of flanking Satt294 indicated that the size variation was due to extra stretch of TTA repeats of the SSR motif in SS2-2, along with $A\longrightarrow$G transversion. In spite of phenotypic differences between the wild type and its hypernodulating mutants, genomic DNA poly-morphisms at microsatellite loci could not control regulation of nodule formation. The cDNA-AFLP method was applied to compare differential display of cDNA between Sinpaldalkong 2 and SS2-2. After isolation and sequence comparison with many AELP fragments, several interesting genes were identified. Northern blot analysis, immunolocalization and/or the yeast two-hybrid system with these genes might provide information on regulation of nodule development in SS2-2.

• Genomics & Informatics
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• v.21 no.4
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• pp.52.1-52.10
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• 2023

Accurate and efficient microbial diagnosis is crucial for effective molecular diagnostics, especially in the field of human healthcare. The gold standard equipment widely employed for detecting specific microorganisms in molecular diagnosis is quantitative real-time polymerase chain reaction (qRT-PCR). However, its limitations in low metagenomic DNA yield samples necessitate exploring alternative approaches. Digital PCR, by quantifying the number of copies of the target sequence, provides absolute quantification results for the bacterial strain. In this study, we compared the diagnostic efficiency of qRT-PCR and digital PCR in detecting a particular bacterial strain (Staphylococcus aureus), focusing on skin-derived DNA samples. Experimentally, specific primer for S. aureus were designed at transcription elongation factor (greA) gene and the target amplicon were cloned and sequenced to validate efficiency of specificity to the greA gene of S. aureus. To quantify the absolute amount of microorganisms present on the skin, the variable region 5 (V5) of the 16S rRNA gene was used, and primers for S. aureus identification were used to relative their amount in the subject's skin. The findings demonstrate the absolute convenience and efficiency of digital PCR in microbial diagnostics. We suggest that the high sensitivity and precise quantification provided by digital PCR could be a promising tool for detecting specific microorganisms, especially in skin-derived DNA samples with low metagenomic DNA yields, and that further research and implementation is needed to improve medical practice and diagnosis.

• KSBB Journal
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• v.16 no.6
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• pp.562-567
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• 2001

Aspergillus niger LK harboring the enantioselective epoxide hydrolase (EHase) activity was isolated, and enantioselectivity of EHase was tested for various racemic aromatic epoxides. The gene encoding epoxide hydrolase was cloned from cDNA library generated by reverse transcriptase-polymerase chain reaction of the isolated total mRNA. Sequence analysis showed that the cloned gene encodes 398 amino acids with a deduced molecular mass of 44.5 kDa. Database comparison of the amino acid sequence reveals that it is similar to fungal EHase, whereas the sequence identity with bacterial EHase is very low. Recombinant expression of the cloned EHase in Escherichia coli BL21 yielded an active EHases, which can offer a potential biocatalyst for the production of chiral epoxides.

• Environmental Analysis Health and Toxicology
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• v.26
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• pp.10.1-10.7
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• 2011

Objects: Chironomus riparius, a non-biting midge (Chironomidae, Diptera), is extensively used as a model organism in aquatic ecotoxicological studies, and considering the potential of C. riparius larvae as a bio-monitoring species, little is known about its genome sequences. This study reports the results of an Expressed Sequence Tags (ESTs) sequencing project conducted on C. riparius larvae using 454 pyrosequencing. Method: To gain a better understanding of C. riparius transcriptome, we generated ESTs database of C.ripairus using pyrosequencing method. Results: Sequencing runs, using normalized cDNA collections from fourth instar larvae, yielded 20,020 expressed sequence tags, which were assembled into 8,565 contigs and 11,455 singletons. Sequence analysis was performed by BlastX search against the National Center for Biotechnology Information (NCBI) nucleotide (nr) and uniprot protein database. Based on the gene ontology classifications, 24% (E-value${\leq}1^{-5}$) of the sequences had known gene functions, 24% had unknown functions and 52% of sequences did not match any known sequences in the existing database. Sequence comparison revealed 81% of the genes have homologous genes among other insects belonging to the order Diptera providing tools for comparative genome analyses. Targeted searches using these annotations identified genes associated with essential metabolic pathways, signaling pathways, detoxification of toxic metabolites and stress response genes of ecotoxicological interest. Conclusions: The results obtained from this study would eventually make ecotoxicogenomics possible in a truly environmentally relevant species, such as, C. riparius.

• Journal of Plant Biotechnology
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• v.47 no.2
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• pp.131-140
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• 2020

Solanum stoloniferum, one of the wild tetraploid Solanum species belonging to the Solanaceae family, is an excellent resource for potato breeding owing to its resistance to several important pathogens. However, the sexual hybridization of S. stoloniferum with S. tuberosum (potato) is hampered due to the sexual incompatibility between the two species. To overcome this and introgress the various novel traits of S. stoloniferum in cultivated potatoes, cell fusion can be performed. The identification of the fusion products is crucial and can be achieved with the aid of molecular markers. In this study, the chloroplast genome sequence of S. stoloniferum was obtained by next-generation sequencing technology, and compared with that of six other Solanum species to identify S. stoloniferum-specific molecular markers. The length of the complete chloroplast genome of S. stoloniferum was found to be 155,567 bp. The structural organization of the chloroplast genome of S. stoloniferum was similar to that of the six other Solanum species studied. Phylogenetic analysis of S. stoloniferum with nine other Solanaceae family members revealed that S. stoloniferum was most closely related to S. berthaultii. Additional comparison of the complete chloroplast genome sequence of S. stoloniferum with that of five Solanum species revealed the presence of six InDels and 39 SNPs specific to S. stoloniferum. Based on these InDels and SNPs, four PCR-based markers were developed to differentiate S. stoloniferum from other Solanum species. These markers will facilitate the selection of fusion products and accelerate potato breeding using S. stoloniferum.

• Microbiology and Biotechnology Letters
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• v.51 no.4
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• pp.500-506
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• 2023

Carbapenem-resistant Klebsiella pneumoniae (CRKP) is emerging as a worldwide public health threat. Recently, Klebsiella pneumoniae carbapenemase-2 (KPC-2)-producing sequence type (ST) 307 was identified main clone of CRKP, and dissemination of ST307 was reported in South Korea. This study examined the molecular characteristic and antimicrobial resistance pattern of 50 CRKP isolated from a tertiary hospital in Daejeon, from March 2020 to December 2021. Epidemiological relationship was analyzed by Multilocus sequence typing (MLST) and antimicrobial susceptibility test was determined using disk-diffusion method. PCR and DNA sequence analysis were performed to identify carbapenemase genes. CRKP infections were significantly more frequent in males and the patients aged ≥ 60 years. Among the 50 CRKP isolates, 46 isolates (92.0%) were multidrug-resistant (MDR), and 44 isolates (88.0%) were carbapenemase-producing K. pneumoniae (CPKP). The major carbapenemase type was KPC-2 (36 isolates, 72.0%) and New Delhi metalloenzyme-1 (NDM-1) and NDM-5 were identified in 7 isolates (14.0%) and 1 isolate (2.0%), respectively. In particular, 88.9% (32/36) of KPC-2-producing K. pneumoniae belonged to ST307, whereas 87.5% (7/8) of NDM-1,-5-producing K. pneumoniae belonged to non-ST307. These results suggest that proper infection control and effective surveillance network need to prevent not olny the spread of ST307, but also the development of non-ST307.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.30 no.1
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• pp.88-94
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• 1997

The mitochondrial DNA (mtDNA) restriction fragment length polymorphi는 (RFLPs) of five populations were analyzed to delineate the stocks of Penaeus chinensis (Osbeck) in the Yellow Sea. Comparison of P. chinensis with P. japonicus to clarify the nucleotide divergence between two species was also carried out. Based on the fragment patterns, three composite haplotypes were analyzed in P. chinensis mtDNA as four naplotypes were in P. japonicus. Most individuals of each P. chinensis population are shared by one dominant haplotype. Another two haplotypes haying variations at the C/a I and hull sites were also distributed evenly in the Korean and Chinese populations. It is suggested that the gene exchange occurring between populations in the Yellow Sea is frequent. Average length of the mtDNA molecule was estimated to be about 16.44 kb in P. chinensis and 16.31 kb in P. japonicus, Sequence divergence (p) of mtDNA between two species estimated by using Upholt's (1977) fomula was $13.7\%$.

• Plant Biotechnology Reports
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• v.3 no.1
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• pp.95-101
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• 2009

Tea leaves are major source of catechins—antioxidant flavonoids. Dihydroflavonol 4-reductase (DFR, EC 1.1.1.219) is one of the important enzymes that catalyzes the reduction of dihydroflavonols to leucoanthocyanins, a key ''late'' step in the biosynthesis of catechins. This manuscript reports characterization of DFR from tea (CsDFR) that comprised 1,413 bp full-length cDNA with ORF of 1,044 bp (115-1,158) and encoding a protein of 347 amino acids. Sequence comparison of CsDFR with earlier reported DFR sequences in a database indicated conservation of 69-87% among amino acid residues. In silico analysis revealed CsDFR to be a membrane-localized protein with a domain (between 16 and 218 amino acids) resembling the NAD-dependent epimerase/dehydratase family. The theoretical molecular weight and isoelectric point of the deduced amino sequence of CsDFR were 38.67 kDa and 6.22, respectively. Upon expression of CsDFR in E. coli, recombinant protein was found to be functional and showed specific activity of 42.85 nmol $min^{-1}$ mg $protein^{-1}$. Expression of CsDFR was maximum in younger rather than older leaves. Expression was down-regulated in response to drought stress and abscisic acid, unaffected by gibberellic acid treatment, but up-regulated in response to wounding, with concomitant modulation of catechins content. This is the first report of functionality of recombinant CsDFR and its expression in tea.

• Journal of Life Science
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• v.17 no.8 s.88
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• pp.1034-1038
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• 2007

Hypericin (HyH) is a substance which is isolated a medicinal herb, Hypericum perforatum L., commonly known as St. John's Wort. Hypericum erectum is a long-lived herb that is distributed in Korea. Cloned HyH genes H. erectum of were conformed by sequencing. The cDNA Hyp-1 sequence has 732 bp with an open reading frame of 567. Thus coding for a protein of 152 amino acid residues. A BLAST re-search using the deduced nucleotide sequences in HyH gene produced significant alignments with the H. perforatum. Sequences in HyH gene showed significant homology with Rubus idaeus putative allergen Rub-i-1 mRNA, Protein sequence comparisons revealed significant homology between Hyp-1 and the phenolic oxidative coupling protein hyp-1 of H. perforatum (98%). Additionally, Hyp-1 showed sig-nificant homology with various other classes of allergens, including Pru-av-1 (62%) from Prunus avium and allergen Bet-vl-Sc3 from Betula pendula (60%). Thus, the result of this study may offer an important information to establish an assay system for chemicals of the herbal medicines for H. erectum as well as H. perforatum.

• Journal of Plant Biotechnology
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• v.4 no.1
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• pp.7-15
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• 2002

Eight cDNAs corresponding to fruit-specific genes were isolated from ripened melon through differential screening. Sequence comparison indicated that six of these cDNAs encoded proteins were previously characterized into aminocyclopropane-1-carboxylate (ACC) oxidase, abscisic acid, stress and ripening inducible (ASR) gene, RINC-H2 zinc finger protein, pyruvate decarboxylase, or polyubiquitin. RFS2 and RFS5 were the same clone encoding polyubiquitin. The other cDNAs showed no significant homology with known protein sequences. The ASR homologue (Asr1) gene was further characterized on the cDNA and genomic structure. The deduced amino acid sequence had similar characteristics to other plant ASR. The Asr1 genomic DNA consisted of 2 exons and 1 intron, which is similar to the structure of other plants ASR genes. The promoter region of the Asr1 gene contained several putative functional cis-elements such as an abscisic acid responsive element (ABRE), an ethylene responsive element (ERE), a C-box or DPBf-1 and 2, Myb binding sites, a low temperature responsive element (LTRE) and a metal responsive element (MRE). The findings imply that these elements may play important roles in the response to plant hormones and environmental stresses in the process of fruit development. The results of this study suggest that the expressions of fruit specific and ripening-related cDNAs are closely associated with the stress response.