• The Plant Pathology Journal
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• v.25 no.3
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• pp.247-255
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• 2009

Root-knot nematode species, such as Meloidogyne hapla, M. incognita, M. arenaria, and M. javanica are the most economically notorious nematode pests, causing serious damage to a variety of crops throughout the world. In this study, DNA sequence analyses were performed on the D3 expansion segment of the 28S gene in the ribosomal DNA in an effort to characterize genetic variations in the three Meloidogyne species obtained from Korea and four species from the United States. Further, PCR-RFLP (Polymerase Chain Reaction-Restriction Fragment Length Polymorphism), SCAR (Sequence Characterized Amplified Region) PCR and RAPD (Randomly Amplified Polymorphic DNA) were also utilized to develop methods for the accurate and rapid species identification of the root-knot nematode species. In the sequence analysis of the D3 expansion segment, only a few nucleotide sequence variations were detected among M. incognita, M. arenaria, and M, javanica, but not M. hapla. As a result of our haplotype analysis, haplotype 5 was shown to be common in M. arenaria, M. incognita, M. javanica, but not in the facultatively parthenogenetic species, M. hapla. PCR-RFLP analysis involving the amplification of the mitochondrial COII and large ribosomal RNA (lrRNA) regions yielded one distinct amplicon for M. hapla at 500 bp, thereby enabling us to distinguish M. hapla from M. incognita, M. arenaria, and M. javanica reproduced via obligate mitotic parthenogenesis. SCAR markers were used to successfully identify the four tested root-knot nematode species. Furthermore, newly attempted RAPD primers for some available root-knot nematodes also provided some species-specific amplification patterns that could also be used to distinguish among root-knot nematode species for quarantine purposes.

• Parasites, Hosts and Diseases
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• v.57 no.1
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• pp.55-60
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• 2019

This study was undertaken to determine the complete mitochondrial DNA sequence and structure of the mitochondrial genome of Spirometra ranarum, and to compare it with those of S. erinaceieuropaei and S. decipiens. The aim of this study was to provide information of the species level taxonomy of Spirometra spp. using the mitochondrial genomes of 3 Spirometra tapeworms. The S. ranarum isolate originated from Myanmar. The mitochondrial genome sequence of S. ranarum was compared with that of S. erinaceieuropaei (GenBank no. KJ599680) and S. decipiens (GenBank no. KJ599679). The complete mtDNA sequence of S. ranarum comprised 13,644 bp. The S. ranarum mt genome contained 36 genes comprising 12 protein-coding genes, 22 tRNAs and 2 rRNAs. The mt genome lacked the atp8 gene, as found for other cestodes. All genes in the S. ranarum mitochondrial genome are transcribed in the same direction and arranged in the same relative position with respect to gene loci as found for S. erinaceieuropaei and S. decipiens mt genomes. The overall nucleotide sequence divergence of 12 protein-coding genes between S. ranarum and S. decipiens differed by 1.5%, and 100% sequence similarity was found in the cox2 and nad6 genes, while the DNA sequence divergence of the cox1, nad1, and nad4 genes of S. ranarum and S. decipiens was 2.2%, 2.1%, and 2.6%, respectively.

• Journal of Practical Agriculture & Fisheries Research
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• v.14 no.1
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• pp.61-83
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• 2012

To identification of fungi that occurs round bale silages, 253 fungal contaminated samples were collected from 2009 to 2011. Total 253 silage samples from Italian ryegrass, sudan grass, rye, corn, barley and oat were analysed. Total 270 strains were purely isolated from contaminated round bale silages. The fungi were identified with morphological characteristics and rDNA sequence analysis. Nineteen species of fungi(Rhizopus sp., Fusarium spp., Coprinus sp., Blastomyces sp., Aureobasidium sp., Polypaecilum sp., Botryoderma sp., Mucor sp., Scytalidium sp., Sphaeropsis sp., Aspergillus spp., Trichocladium sp., Humicola sp., Staphylotrichum sp., Periconia sp., Verticillium sp., Diplococcium sp., Penicillium spp. and Trichoderma spp.) were identified by morphological characteristics. On the other hand, fungi isolated from silage were identified to Acremonium strictum, Aspergillus tubingensis, Bionectria ochroleuca, Dipodascaceae sp., Fusarium proliferatum, Fusarium oxysporum, Fusrium solani, Gelasinospora reticulata, Gibberella moniliformis, Gibberella zeae, Nectria mauritiicola, Penicillium paneum, Pseudallecheria boydii, Schizophyllum commune, Scopulariopsis brevicaulis and Simplicillium lamellicola by rDNA sequence analysis. Penicillium sp. and Trichoderma sp., were isolated 74 and 64 strains, respectively. Humicola sp., Aspergillus sp., Coprinus sp., and Fusarium spp. were identified 10 to 30 strains. Most fungi were isolated together with more than one species in a sample looked like one species with the naked eyes.

• Proceedings of the Microbiological Society of Korea Conference
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• 2002.10a
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• pp.15-21
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• 2002

Terminal-restriction fragment length polymorphism (T-RFLP) analysis has been optimized by using in vitro model community composed of genomic DNAs of known bacterial strains and has been applied to assess the bacterial community structure in marine sediments. The specific fluorescence-labeled terminal restriction fragments (T-RFs) between 39 and 839 base long specifying each strain were precisely measured for known bacterial strains. The addition of a co-solvent (dimethylsulfoxide or glycerol) into PCR reactions has reduced differential PCR amplification. Comparative bacterial community structure was investigated for pristine and polluted sediments. A complex T-RFLP pattern showing complex bacterial community structure was obtained in the pristine sediment, whereas simple T-RFLP pattern (low bacterial diversity) was shown in polluted sediments where caged aquaculture has been conducted for several years. The results of T-RFLP analysis were compared with that of cloning and sequencing 16S rDNA clones from the same sediments. Sequence analysis of 16S rDNA clones (72) of the pristine sediment revealed a diverse collection of lineages, largely of the class Proteobacteria ($6\%$ alpha subdivision, $46\%$ gamma subdivision, $13\%$ delta subdivision, and $3\%$ epsilon subdivision), Nitrospina $(8\%)$, high G+C gram positive $(8\%)$, Verrucomicrobia $(7\%)$, and Planctomycetes $(6\%)$. In the contaminated sediments, 17 $(59\%)$ of the 16S rDNA clones (29) were related to Campylobacter and symbiont of Rimicaris exoculata belonging to epsilon subdivision of Proteobacteria. The results obtained indicated that T-RFLP analysis is a rapid and precise technique for comparative bacterial community analysis.

• The Plant Pathology Journal
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• v.18 no.3
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• pp.109-114
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• 2002

To evaluate the phylogenetic and taxonomic status of the phytoplasmas associated with water dropwort (Oenanthe javanica DC) disease in Korea and Japan, their 16S rDNA was analyzed. DNAs extracted from water dropworts collected in Korea (Kyongnam province) and Japan (Chiba prefecture) affected by witches' broom and yellows were subjected to PCR using phytoplasma-specific primers, which amplified a 1.4-kbp fragment that included the 16S rDNA. Phytoplasmas were characterized by RFLP analysis using AluI, HaeIII, HhaI, KpnI, MseI, and RsaI restriction enzymes and by sequence analysis of the PCR products. The mater dropwort witches'broom (WDWB) and water dropwort yellows (WDY) 16S rDNA sequences were identical and closely related to onion yellows (OY, 99.9% identity), which belong to the aster yellows (AY) 16S-subgroup. However, the KpnI RFLP analyses clearly distinguished the WDY and WDWB phytoplasmas from the OY phytoplasma. The phylogenetic analysis based on 16S rDNA showed that WDWE and WDY phytoplasmas are members of a relatively homogeneous group that evolved from a common ancestor.

• Journal of Microbiology and Biotechnology
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• v.12 no.5
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• pp.773-779
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• 2002

A gene (hap) encoding aminopeptidase from the chromosomal DNA of Bacillus licheniformis was cloned. The gene is 1,347 bp long and encodes a 449 amino acid preproprotein with a major mature region of 401 amino acids (calculated molecular mass 43,241 Da). N-Terminal sequence of the purified protein revealed a potential presence of N-terminal propeptide. The deduced primary amino acid sequence and the mass analysis of the purified protein suggested that a C-terminal peptide YSSVAQ was also cleaved off by a possible endogeneous protease. Tho amino acid sequence displayed 58% identity with that of the aminopeptidase from alkaliphilic Bacillus halodurans. This bacterial enzyme was overexpressed in recombinant Escherichia coli and Bacillus subtilis cells. Clones containing the intact hap gene, including its own promoter and signal sequence, gave rise to the synthesis of extracellular and thrmostable enzyme by B. subtilis transformants. The secreted protein exhibited the same biochemical properties and the similar apparent molecular mass as the B. lichenzyormis original enzyme.

• Fisheries and Aquatic Sciences
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• v.13 no.4
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• pp.272-277
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• 2010

Seagrasses are marine angiosperms of ecological importance in providing shelter and food to aquatic species as well as maintaining the carbon cycle on earth. Phyllospadix iwatensis is a seagrass of the family Zosteraceae and is distributed along the eastern coast of Korea. The nucleotide sequences of P. iwatensis nuclear genes encoding 18S ribosomal RNA (rRNA) and internal transcribed spacer-1 (ITS-1) were determined for molecular phylogenetic analysis. Genomic DNA was isolated from P. iwatensis and used for PCR amplification of 18S rRNA and ITS-1. Examination of the 18S rRNA sequence of P. iwatensis showed a close (99% similarity) relationship to Zostera noltii, another genus of Zosteraceae, but a distant (84% similarity) evolutionary relationship to other macroalgal Laminariales species. Further discrepancies found in ITS-1 nucleotide sequences between closely related species indicate that the sequence information could be used for species identification.

• Korean Journal of Veterinary Research
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• v.45 no.3
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• pp.351-357
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• 2005

Here I describe a multiplex allele specific PCR-based approach for the rapid detection between Hanwoo and Holstein meat associated with Melanocortin 1 receptor (MC1R) gene. Specific and universal oligonucleotide primers were used in combination to detect the presence of a single nucleotide polymorphism within the bovine MC1R DNA sequence. The presence of the bovine MC1R gene is indicated by the production of a single control PCR product, whilst positive samples generate an alternative smaller specific product over the same region. The mutations in MC1R104 codon revealed depending on the presence or absence of an indicative fragment amplified from the wild-type allele of this codon. As little as 0.39 ng and 1.56 ng of genomic DNA of Hanwoo and Holstein could be detected by MAS-PCR assay, respectively. This technique, which is widely used in human genetic screening, provides a reliable and sensitive result that has not been documented for the identification of bovine coat color. The MAS-PCR assay approach was proven to be useful in complementing routine beef DNA analysis for differentiation of these MC1R variants and it would facilitate the screening of deceiving sales of Holstein meat in the butcher shop.

• Research in Plant Disease
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• v.18 no.2
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• pp.73-79
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• 2012

Phytophthora katsurae is a fungal pathogen responsible for chestnut ink disease. We designed two duplex primer sets (SOPC 1F/1R+KatI 3F/5R, SOPC 1-1F/1-1R+KatI 3F/5R) to detect P. katsurae. SOPC 1F/1R and SOPC 1-1F/1-1R primer pairs were designed for sequence characteristic amplification regions (SCAR) marker, and KatI 3F/5R primer pair was used for P. katsurae-specific primer designed from internal transcribed spacer (ITS) region. To assess the sensitivity of duplex PCR, genomic DNA was serially diluted 10-fold to make the final concentrations from 1 mg/ml to 1 ng/ml. The sensitivity for two primer sets were 1 ${\mu}g/ml$ and 100 ng/ml, respectively. To find detection limits for zoospores of P. katsurae, each zoospore suspension was serially diluted 10-fold to make the final concentrations from $1{\times}10^6$ to $1{\times}10^2$ cells/ml, and then DNA was extracted. The limits of detection for all of two primer sets were $1{\times}10^5$ cells/ml. All of two primer sets were specific to P. katsurae in PCR detection and did not produce any P. katsurae-specific PCR amplicons from other 16 Phytophthora species used as the control. This study shows that duplex PCR using two primer sets might be a useful tool for rapid and efficient detection of P. katsurae.

• Journal of Microbiology and Biotechnology
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• v.27 no.3
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• pp.535-541
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• 2017

Two Gram-staining-negative, red-pinkish, coccus-shaped, non-motile, and aerobic bacterial strains, designated $Ant21^T$ and Ant22, were isolated from the Antarctic coastal sea water. Strains $Ant21^T$ and Ant22 showed UVC and gamma radiation resistance. Phylogenetic analyses based on 16S rRNA gene sequences determined that these strains belong to the genus Deinococcus. Through the analyses of the 16S rRNA gene sequences, strains $Ant21^T$ and Ant22 were found to have 97.7% and 97.8% similarity to Deinococcus marmoris DSM $12784^T$ and 97.0% and 97.2% similarity to Deinococcus saxicola AA-$1444^T$, respectively. The sequence similarity with the type strains of other Deinococcus species was less than 96.9% for both strains. Strains $Ant21^T$ and Ant22 shared relatively high 16S rRNA gene sequence similarity (99.3%) and had a closely related DNA reassociation value of $84{\pm}0.5%$. Meanwhile, they showed a low level of DNA-DNA hybridization (<30%) with other closely related species of the genus Deinococcus. The two strains also showed typical chemotaxonomic features for the genus Deinococcus, in terms of the major polar lipid (phosphoglycolipid) and the major fatty acids ($C_{16:0}$, $C_{16:1}$ ${\omega}6c/{\omega}7c$, $iso-C_{17:0}$, and $iso-C_{15:0}$). They grew at temperatures between $4^{\circ}C$ and $30^{\circ}C$ and at pH values of 6.0-8.0. Based on the physiological characteristics, the 16S rRNA gene sequence analysis results, and the low DNA-DNA reassociation level with Deionococcus marmoris, strains $Ant21^T$ ($=KEMB\;9004-167^T$ $=JCM\;31436^T$) and Ant22 (KEMB 9004-168 =JCM 31437) represent novel species belonging to the genus Deinococcus, for which the name Deinococcus rubrus is proposed.