• Proceedings of the Plant Resources Society of Korea Conference
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• 2018.10a
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• pp.74-74
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• 2018

Little millet (Panicum sumatrense) is well known for its salt and drought stress tolerance and high nutritional value, but very limited knowledge of genetic variation and genomic information is available. This study was to develop highly polymorphic EST-SSR markers based on cross-species transferability of derived SSRs from switchgrass EST databases and characterize newly developed EST - SSRs to better understand the genetic diversity of collected 37 germplasm accessions of little millet. A total of 779 primer pairs were designed from the 22,961 EST sequences of switchgrass (Pancium virgatum), of which 48 EST - SSR markers were developed based on the trials of transferability of these primers in little millet. The EST - SSR amplicons showed reproducible single band polymorphism and produced a total of 160 alleles with an average of 3.3 alleles per locus in 37 accessions of little millet. T he average values of expected and observed heterozygosities were 0.266 and 0.123, respectively. T he polymorphic information content (PIC) values were observed in range of 0.026 to 0.549 with an average of 0.240. The genetic relatedness among the little millet accessions was evaluated by neighbor-joining dendrogram, which grouped all accessions into two distinct groups. The validation thus demonstrated the utility of the switchgrass EST - SSR markers in assessing genomic relationships in little millet. T he findings from this study could be useful for designing strategies for the identification of diverse germplasm for conservation and future molecular breeding programs for little millet.

• Bulletin of the Korean Chemical Society
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• v.32 no.6
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• pp.1925-1930
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• 2011

Psychrophilic bacteria have acquired cold-resistance in order to protect themselves against freezing temperatures, which would otherwise be lethal. DnaK/DnaJ/GrpE systems are molecular chaperones which facilitate proper folding of newly synthesized proteins. Efficient folding processes are of great importance especially in a cold environment, such as the Arctic. In order to understand the protection mechanisms of psychrophilic bacteria against cold temperatures, we have explored a genome of KOPRI22215, tentatively identified as Psychromonas arctica, whose genome sequence has not yet been discovered. With an aim of searching for a coding gene of DnaK from KOPRI22215, we have applied a series of polymerase chain reactions (PCR) with homologous primers designed from other Psychromonas species and LA PCR in vitro cloning. 1917 bp complete coding sequence of dnaK from KOPRI22215 was identified including upstream promoter sites. Recombinant plasmids to overexpress PaDnaK along with EcDnaK (DnaK of E. coli) were then constructed in pAED4 vector and the pET-based system to induce PaDnaK expression by IPTG. Characterization assays of expressed PaDnaK were carried out by measuring survival rates upon 4 day incubation at 4 $^{\circ}C$: a refolding assay as molecular chaperone, and ATPase assay for functional activity. Taking account of all the data together, we conclude that PaDnaK was identified, successfully expressed, and found to be more efficient in providing cold-resistance for bacterial cells.

• Korean Journal of Ecology and Environment
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• v.54 no.3
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• pp.190-198
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• 2021

Human Aichivirus (Aichivirus A; AiV-A) is a positive-sense single-strand RNA non-enveloped virus that has been detected worldwide in various water environments including sewage, river, surface, and ground over the past decade. To develop a method with excellent sensitivity and specificity for AiV-A diagnosis from water environments such as groundwater, a combination capable of reverse transcription (RT)-nested polymerase chain reaction (PCR) was developed based on existing reported and newly designed primers. A selective method was applied to evaluate domestic drinking groundwater samples. Thus, a procedure was devised to select and subsequently identify RT-nested PCR primer sets that can successfully detect and identify AiV-A from groundwater samples. The findings will contribute to developing a better monitoring system to detect AiV-A contamination in water environments such as groundwater.

• Korean Journal of Microbiology
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• v.49 no.3
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• pp.270-274
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• 2013

The bumblebee, Bombus terrestris, has played an important role as one of the alternative pollinators since the outbreak of honeybee collapse disorder. Recently, pathogens and parasites such as viruses, bacteria and mites, which affect the life span and fecundity of their host, have been discovered in B. terristris. In order to detect the microsporidian pathogen, Nosema spp. in the field populations of B. terristris, we collected adults and isolated their genomic DNA for diagnostic PCR. The PCR primers specific for Nosema spp. were newly designed and applied to gene amplification for cloning. Only small subunit ribosomal RNA (SSU rRNA) gene of N. ceranae was successfully amplified among examined genes and sequenced, which indicates that N. ceranae mainly infects the examined field population of B. terristris. To detect of SSU rRNA gene, two regions of SSU rRNA gene were selected by primary PCR analysis and further analyzed in quantitative real-time PCR (qRT-PCR). The qRT-PCR analysis demonstrated that SSU rRNA of N. ceranae was detected at concentration as low as $0.85ng/{\mu}l$ genomic DNA. This result suggests that the detection via qRT-PCR can be applied for the rapid and sensitive diagnosis of N. ceranae infection in the field population as well as risk assessment of B. terristris.

• Journal of Microbiology and Biotechnology
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• v.19 no.5
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• pp.525-529
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• 2009

Cytolethal distending toxins (CDTs) represent an emerging family of newly described bacterial products that are produced by a number of pathogens. The genes encoding these toxins have been identified as a cluster of three adjacent genes, cdtA, cdtB, and cdtC, plus 5 cdt genetic variants, designated as cdt-I, cdt-II, cdt-III, cdt-IV, and cdt-V, have been identified to date. In this study, a general multiplex PCR system designed to detect Escherichia coli cdts was applied to investigate the presence of cdt genes among isolates. As a result, among 366 E. coli strains, 2.7% were found to carry the cdtB gene. In addition, the use of type-specific primers revealed the presence of cdt-I, cdtIV, and cdt-V types of the cdt gene, yet no cdt-II or cdt-III strains. The presence of other virulence genes (stxl, stx2, eae, bfp, espA, espB, and espD) was also investigated using a PCR assay. Among the 10 cdtB gene-positive strains, 8 were identified as COT-producing typical enteropathogenic E. coli (EPEC) strains ($eae^+$, $bfp^+$), whereas 2 were identified as CDT-producing atypical EPEC strains ($eae^+$, $bfp^-$). When comparing the cytotoxic activity of the CDT-producing typical and atypical EPEC strains, the CDT-producing atypical EPEC strains appeared to be less toxic than the CDT-producing typical EPEC strains.

• Journal of Korean Society of Forest Science
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• v.108 no.3
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• pp.302-310
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• 2019

Precise and fast identification of crop cultivars is essential for efficient breeding and plant breeders' rights. Traditional methods for identification of jujube cultivars are based on the evaluation of morphological characteristics. However, due to time constraints and environmental influences, it is difficult to distinguish cultivars using only morphological traits. In this study, we cloned fragments from improved inter simple sequence repeats (ISSR) analysis, and developed stably diagnostic sequence-characterized amplified region (SCAR) markers. The specific ISSR bands of jujube cultivars from Dalizao and Boeundaechu were purified, cloned, and sequenced. As a result, four clones labeled 827Dalizao550, 827Boeun750, 846Boeun700, and 847Dalizao850 were identified. In order to investigate whether they were specific for the jujube cultivar, four pairs of SCAR primers were then designed and polymerase chain reaction (PCR) amplifications were conducted to analyze 32 samples, including jujube and sour jujube. In the PCR amplification of the 827Dalizao550 SCAR marker, the specific bands with 550 bp were amplified in six samples (Dalizao, Sandonglizao, Dongzao, Yuanlin No. 2, Suanzao 2, Suanzao 4), but unexpected bands (490 bp) were amplified in the others. Moreover, in the PCR amplification of the 847Dalizao850 SCAR marker, the specific bands with 850 bp were found in three samples (Dalizao, Sandonglizao, and Dongzao) and 900 bp unexpected bands were amplified in five samples (Pozao, Suanzao 1, Suanzao 2, Suanzao 3, Suanzao 4). These results showed that newly developed markers could be useful as a fast and reliable tool to identify jujube cultivars. However, further identification of polymorphic information and the development of SCAR markers are required for the identification of more diverse cultivars.

• Clinical and Experimental Pediatrics
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• v.45 no.10
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• pp.1263-1272
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• 2002

Purpose : Rett syndrome(RTT) is an X-linked dominant neurodevelopmental disorder affecting 1 per 10,000-15,000 female births worldwide. It was initially described by Andreas Rett in 1966. RTT involves developmental regression characterized stereotypic hand movements, tremors, gait apraxia, seizures, deceleration of head growth after the age of 6-18 months. The disease-causing gene was identified as MECP2 on chromosome Xq28. We carried out mutational analysis of MECP2 genes in RTT patients. Methods : Whole blood(5 cc) of 34 sporadic RTT patients was collected in EDTA-anticoagulated tubes. Genomic DNA was extracted from peripheral blood using the E.Z.N.A. blood DNA kit. Four exons of the MECP2 gene were amplified by PCR in 34 Korean with RTT. We carried out PCR divided the exon three into two parts and the exon four into five parts. Primer sequences designed by Amir et al. in 1999 were almost used(AF030876). Sequencing primers used were the same as PCR. DNA sequencing reactions were performed using an ABI 377 DNA sequencer and ABI PRISM dye terminator cycle sequencing reaction kit(Perkin-elmer). The results were compared with the normal DNA sequence(X99686). To confirm the change of sequence on novel mutations, RFLP analysis was performed. Results : The MECP2 mutations were detected in 23(67.6%) of the 34 patients. The mutations consisted of 12 different types including nine missense and three nonsense mutations. Of these, three (L100V, G161E and T311M) mutations were newly identified. Most of the mutations discovered are located within MBD(39.1%) and TRD(39.1%). In this study, three(T158M, R270X, R306C) mutations were identified high frequency. Conclusion : MECP2 gene was also an important cause of Korean RTT patients. MECP2 gene study is an important tool for diagnosis of Korean RTT patients.