• The Korean Journal of Mycology
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• v.25 no.3 s.82
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• pp.176-190
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• 1997

Sixty-three isolates of Lentinus edodes obtained from Korea were used to assess the genetic similarity by isozyme polymorphisms and random amplified polymorphic DNA patterns. The activities of esterase, peroxidase and acid phosphatase displayed 10, 7 and 3 distinct isozyme patterns, respectively. By combining the isozyme patterns obtained with the 3 enzymes, every isolate showed its own distinct electrophoretic phenotypes. A distance matrix calculated between all pairs of 63 electrophoretic phenotypes based on the presence or abscence of isozyme bands were analyzed by the group-average method. Results of the cluster analysis assinged the 63 phenotypes into six major groups. In the analysis of random amplified polymorphic DNA patterns, all isolates of Lentinus edodes were devided into five RAPD groups.

• Journal of Life Science
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• v.28 no.5
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• pp.555-562
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• 2018

Swine influenza virus (SIV) causes one of the most common diseases of the pig population, and its subtypes are determined by hemagglutinin (HA) and neuraminidase (NA). Recently, the SIV subtype diagnosis has been developed. The method using antigen-antibody reaction rather than PCR was mainly used because of the large change in the ribonucleotide sequences of SIV. Here, we have developed 10 diagnostic primer sets through multi-nucleotide sequences alignment of spreaded SIV since 2008 in Korea and then optimized the reaction of the one-step RT-PCR for rapid determination of SIV subtype. In addition, specific primers were designed to early determine the pandemic SIV by detecting unique M sequences proven in highly infectious and virulent subtypes of the influenza H1N1 (pH1N1). Here, some of the SIVs spread in Korea from 2008 to 2014 have been tested to determine the subtypes and pandemic potential of SIV. All diagnostic primer sets were found to be able to accurately determine the SIV subtype and to detect the pandemic SIV. In conclusion, it was confirmed that the optimized one-step RT-PCR analysis using these primer sets is useful for rapid diagnosis of SIV subtypes. These results can be used for development of SIV subtype diagnostic kit to early detect before virulent SIV spreads do.

• Mycobiology
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• v.48 no.4
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• pp.313-320
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• 2020

In Pleurotus sp., green mold, which is considered a major epidemic, is caused by several Trichoderma species. To develop a rapid molecular marker specific for Trichoderma spp. that potentially cause green mold, eleven Trichoderma species were collected from mushroom farms and the Korean Agricultural Culture Collection (KACC). A dominant fungal isolate from a green mold-infected substrate was identified as Trichoderma pleuroticola based on the sequences of its internal transcribed spacer (ITS) and translation elongation factor 1-α (tef1) genes. In artificial inoculation tests, all Trichoderma spp., including T. atroviride, T. cf. virens, T. citrinoviride, T. harzianum, T. koningii, T. longibrachiatum, T. pleurotum, and T. pleuroticola, showed pathogenicity to some extent, and the observed symptoms were soaked mycelia with a red-brown pigment and retarded mycelium regeneration. A molecular marker was developed for the rapid detection of wide range of Trichoderma spp. based on the DNA sequence alignment of the ITS1 and ITS2 regions of Trichoderma spp. The developed primer set detected only Trichoderma spp., and no cross reactivity with edible mushrooms was observed. The detection limits for the PCR assay of T. harzianum (KACC40558), T. pleurotum (KACC44537), and T. pleuroticola (CAF-TP3) were found to be 500, 50, and 5 fg, respectively, and the detection limit for the pathogen-to-host ratio was approximately 1:10,000 (wt/wt).

• Animal cells and systems
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• v.1 no.2
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• pp.363-370
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• 1997

RNase MRP is a ribonucleoprotein complex with a site-specific endonuclease activity. Its original substrate for cleavage is the small mitochondrial RNA near the mitochondrial DNA replication origin, thus it was proposed to generate the primer for mtDNA replication. Recently, it has been shown to have another substrate in the nucleus, such as pre-S.8S ribosomal RNA in nucleolus. The gene for the RNA component of RNase MRP (MRP RNA) was found to be encoded by the nucleus genome, suggesting an interesting intracellular trafficking of MRP RNA to both mitochondria and nucleolus after transcription in nucleus. In this study, genomic DNA encoding MRP RNA was microinjected into the nucleus of Xenopus oocytes, to analyze promoter regions involved in the transcription. It showed that the proximal sequence element and TATA box are important for basal level transcription; octamer motif and Sp1 binding sites are for elevated level transcription. Most of Xenopus MRP RNA was exported out to the cytoplasm following transcription in the nucleus. Utilizing various hybrid constructs, export of MRP RNA was found to be regulated by the promoter and the 5' half of the coding region of the gene. Interestingly, the transcription in nucleus seems to be coupled to the export of MRP RNA to cytoplasm. Intracellular transport of injected MRP RNA can be easily visualized by whole-mount in situ hybridization following microinjection; it also shows possible intra-nuclear sites for transcription and export of MRP RNA.

• Biomedical Science Letters
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• v.23 no.3
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• pp.223-229
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• 2017

Fungal infections by human pathogenic fungi are increasing globally in elderly, children and immune suppressed or deficient patients. Aspergillus fumigatus is one of the well-known pathogenic fungi and causes aspergilloses in human world widely. However, current identification and classification methods based on its phenotypic characteristics still have limitations. Therefore, currently, molecular biological tools using their DNA sequences are used for genotype identification and classification. In the present study, in order to analyze genetic variations of A. fumigatus clinical isolates, a total of six housekeeping genes were amplified by PCR using specific primer pairs and multi-locus sequence typing (MLST) assay. Results from phylogenetic tree analysis showed that most A. fumigatus strains (88.9%) from respiratory specimens were classified into cluster A and B, and approximately half of A. fumigatus strains (46%) from non-respiratory specimens were classified into cluster C and D. Although the sample size was limited, genetic characteristics of A. fumigatus clinical isolates according to their origins were very similar and well-correlated with other clinical data.

• Biomedical Science Letters
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• v.13 no.4
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• pp.319-324
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• 2007

One-tube nested polymerase chain reaction (PCR) was evaluated for its efficacy in detecting Mycobacterium leprae in biopsy samples from leprosy patients. Primers were derived from the M leprae-specific element (RLEP) sequences which yield a 230 bp fragment. The specificity and the sensitivity of the one-tube nested PCR were compared with those of single PCR for detecting M leprae. The results showed that the one-tube nested PCR was about 100 times more sensitive than that of the single indicating the one-tube nested primer sets developed in this study can be an effective screening tool for the detection of M leprae in clinical diagnostic laboratories.

• The Journal of Korean Society of Virology
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• v.29 no.1
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• pp.23-31
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• 1999

Hantavirus is a genus of the Bunyaviridae family causing two serious diseases, hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). Puumala virus is a member of hantavirus originally found in Europe, and its natural reservoir is Clethrionomys glareolus. It is also associated with the human disease nephropathia epidemica, a milder form of HFRS. To identify the hantaviruses in bats, bats were collected from Jeong-Sun, Won-Joo, Chung-Ju and Hwa-Cheon area in Korea, and nested RT-PCR was performed with serotype specific primer from M segment. Interestingly, Puumala virus was detected in bats (Rhinolophus ferrum-equinum) only from Won-Joo. The 327 bp nested RT-PCR product, was sequenced. The sequence database search indicates that the sequence is homologous to the published sequence of Puumala viruses. The sequence similarities were ranged from 71% to 97%. The highest sequence similarity was 97% with Puumala virus Vranicam strain, and the lowest was 71% with Puumala virus K27 isolate. Puumala virus Vranicam strain was isolated from a bank vole (Clethrionomys glareolus) in Bosnia-Hercegovina. Puumala virus K27 was isolated from human in Russia. This analysis confirms that bats (Rhinolophus ferrum-equinum) in Korea are natural reservoir of Puumala virus.

• The Plant Pathology Journal
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• v.20 no.2
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• pp.135-141
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• 2004

A filamentous virus was isolated from taro (Colocasia esculenta Schott) showing mosaic and chlorotic feather-ing symptoms in Chuncheon, Gangwon province in 2002. Based on ELISA, its appearance in electron microscope, serological relationships, and RT-PCR using specific primer and nucleotide sequence analysis of the CP gene, the isolated virus was identified as Dasheen mosaic virus (DsMV) and designated as Korean isolated (DsMV-Kr). DsMV was not serologically related to Zantedeschia mosaic virus (ZaMV), which has been reported to infect an Araceae plants. Since the coat protein revealed electrophoretic heterogeneity, about 42 kDa, 39 kDa and 31 kDa by SDS-PAGE, an improved purification method was established for the production of antisera against DsMV-Kr. The purification method used in this study may be effectively applied to the purification of other filamentous viruses.

• The Plant Pathology Journal
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• v.22 no.1
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• pp.36-45
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• 2006

Twenty isolates of Didymella bryoniae were isolated from infected cucurbit plants in various growing areas of southern Korea in 2001 and 2002. Random Amplified Polymorphic DNA (RAPD) group [RG] I of D. bryoniae was more virulent than RG IV to watermelon. Virulence of the RG I isolate was strong to moderate to cucumber, whereas that of the RG IV varied from strong, moderate to weak. Two hundred seventy-three amplified fragments were produced with 40 primers, and were analyzed by a cluster analysis using UPGMA method with an arithmetic average program of NTSYSPC. At the distance level of 0.7, two major genomic DNA RAPD groups were differentiated among 20 isolates. The RG I included 7 isolates from watermelon and one isolate from melon, whereas the RG IV included 12 isolates from squash, cucumber, watermelon and melon. Amplification of internal transcribed spacer (ITS) region and small subunit rRNA region from the 20 isolates yielded respectively a single fragment. Restriction pattern with 12 restriction enzymes was identical for all isolates tested, suggesting that variation in the ITS and small subunit within the D. bryoniae were low. Amplification of the genomic DNAs of the tested isolates with the sequence characterized amplified regions (SCAR) primer RG IF-RG IR specific for RG I group resulted in a single band of 650bp fragment for 8 isolates out of the 20 isolates. Therefore, these 8 isolates could be assigned into RG I. The same experiments done with RG IIF-RG IIR resulted in no amplified PCR product for the 20 isolates tested. An about 1.4 kb-fragment amplified from the RG IV isolates was specifically hybridized with PCR fragments amplified from genomic DNAs of the RG IV isolates only, suggesting that this PCR product could be used for discriminating the RG IV isolates from the RG I isolates as well other fungal species.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.48 no.5
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• pp.731-738
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• 2015

Nematode infection in the epithelial tissue of cultured rockfish Sebastes schlegeli was first reported in 2012. Since then, nematode infections have caused serious economic losses in rockfish aquaculture on the west coast of Korea. Taxonomic and life cycle information for this parasite are currently unknown. In this study, 18S rRNA and cytochrome c oxidase subunit I (COI) genes were used for molecular identification and polymerase chain reaction (PCR) to detect the invisible stages of this parasite. Nucleotide sequences of the 18S rRNA of the rockfish nematode showed 98% identity with that of Philometra morii. Therefore, this rockfish nematode was classified to the Philometridae family. However, we could not identify it to genus level using 18S rRNA. Its COI nucleotide sequences shared 85% and 82% identities with those of Bursaphelenchus sinensis and Philometra overstreeti, respectively. In addition, two gene-specific primer sets were designed based on the 18S rRNA gene to detect the intermediate host and nematode larvae. These primers were specific to this rockfish nematode without cross-reacting to other pathogens. The detection limit of the PCR assay using these primers was 1,000 copies of nematoda plasmid DNA. Therefore, the PCR assay described here is suitable for the detection of nematode DNA within rockfish. In addition, this PCR assay could be used to detect nematode larvae and the intermediate host.