• Fisheries and Aquatic Sciences
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• v.15 no.2
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• pp.157-162
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• 2012

To analyze nucleotide sequence encoding internal transcribed spacer (ITS) regions specific to the Laminariaceae family, genomic DNA was isolated from six brown algae species distributed along the east coast of Korea. These included three species from the Laminariaceae family (Agarum clathratum Dumortier, Costaria costata [C. Agardh] Saunders, and Saccharina japonica Areschoug) and two species from the Alariaceae family (Undaria pinnatifida [Harvey] Suringer and Ecklonia cava Kjellman), both in the order Laminariales, and one species from the family Sargassaceae in the order Fucales (Sargassum serratifolium). Based on a sequence analysis of ITS-1 and ITS-2 for A. clathratum, C. costata, and E. cava, oligonucleotides were designed from the regions that showed sequence conservation in Laminariaceae. Following polymerase chain reaction using three sets of primers, amplification of ITS-1 and ITS-2 was detected in reactions using genomic DNA isolated from the species belonging to Laminariaceae, but not from the species belonging to the other families. The results indicate that this method can be used for the detection and identification of Laminariaceae species.

• Mycobiology
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• v.50 no.5
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• pp.382-388
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• 2022

White mold (or Sclerotinia stem rot), caused by Sclerotinia species, is a major air, soil, or seed-transmitted disease affecting numerous crops and wild plants. Microscopic or culture-based methods currently available for their detection and identification are time-consuming, laborious, and often erroneous. Therefore, we developed a multiplex quantitative PCR (qPCR) assay for the discrimination, detection, and quantification of DNA collected from each of the three economically relevant Sclerotinia species, namely, S. sclerotiorum, S. minor, and S. nivalis. TaqMan primer/probe combinations specific for each Sclerotinia species were designed based on the gene sequences encoding aspartyl protease. High specificity and sensitivity of each probe were confirmed for sclerotium and soil samples, as well as pure cultures, using simplex and multiplex qPCRs. This multiplex assay could be helpful in detecting and quantifying specific species of Sclerotinia, and therefore, may be valuable for disease diagnosis, forecasting, and management.

• Korean Journal of Ecology and Environment
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• v.53 no.3
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• pp.286-294
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• 2020

The Chironomus(Diptera: Chironomidae) is a freshwater benthic invertebrate that is an important indicator organism used for environmental pollution and water quality monitoring. In this study, we performed morphological classification and genetic species identification using the cytochrome c oxidase subunit I (COI) gene on mitochondrial DNA for an accurate species classification of Chironomus larvae found in tap water purification plants in Incheon, Korea. Twenty larvae in six water purification plants consist of four species, including twelve Chironomus kiiensis, six Chironomus flaviplumus, one Chironomus dorsalis, and one Polypedilum yongsanensis (not included Genus Chironomus). Morphological characteristics of each larvae were identified based on the head capsule, the mentum, the mandible, the antenna, and the claw. Based on the COI sequences of 21 individuals of 17 Chironomus species registered in NCBI Genbank, phylogenetic analysis indicated that the 20 individuals investigated in this study consist of the same clade with corresponding species of the high homology (99~100%) including C. kiiensis, C. flaviplumus, C. dorsalis, and P. yongsanensis. These results will be used as main classification indicator for monitoring freshwater ecosystems by providing integrated morphological and genetic information for the species identification of Korean Chironomus.

• ALGAE
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• v.35 no.3
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• pp.293-301
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• 2020

The applications of DNA barcoding have a wide range of uses, such as in taxonomic studies to help elucidate cryptic species and phylogenetic relationships and analyzing environmental samples for biodiversity monitoring and conservation assessments of species. After obtaining the DNA barcode sequences, sequence similarity-based homology analysis is commonly used. This means that the obtained barcode sequences are compared to the DNA barcode reference databases. This bioinformatic analysis necessarily implies that the overall quantity and quality of the reference databases must be stringently monitored to not have an adverse impact on the accuracy of species identification. With the development of next-generation sequencing techniques, a noticeably large number of DNA barcode sequences have been produced and are stored in online databases, but their degree of validity, accuracy, and reliability have not been extensively investigated. In this study, we investigated the extent to which the amount and types of erroneous barcode sequences were deposited in publicly accessible databases. Over 4.1 million sequences were investigated in three largescale DNA barcode databases (NCBI GenBank, Barcode of Life Data System [BOLD], and Protist Ribosomal Reference database [PR2]) for four major DNA barcodes (cytochrome c oxidase subunit 1 [COI], internal transcribed spacer [ITS], ribulose bisphosphate carboxylase large chain [rbcL], and 18S ribosomal RNA [18S rRNA]); approximately 2% of erroneous barcode sequences were found and their taxonomic distributions were uneven. Consequently, our present findings provide compelling evidence of data quality problems along with insufficient and unreliable annotation of taxonomic data in DNA barcode databases. Therefore, we suggest that if ambiguous taxa are presented during barcoding analysis, further validation with other DNA barcode loci or morphological characters should be mandated.

• Journal of Microbiology and Biotechnology
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• v.16 no.2
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• pp.256-263
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• 2006

In the model legume Medicago truncatula, two mutants, sickle and sunn, exhibit morphologically and genetically distinct hypernodulation phenotypes. However, efforts to isolate the single recessive and single semidominant genes for sickle and sunn, respectively, by map-based cloning have so far been unsuccessful, partly due to the absence of clones that enable walks from linked marker positions. To help resolve these difficulties, a new bacterial artificial chromosome (BAC) library was constructed using BamHI-digested genomic fragments. A total of 23,808 clones were collected from ligation mixtures prepared with double-size-selected high-molecular-weight DNA. The average insert size was 116 kb based on an analysis of 88 randomly selected clones using NotI digestion and pulsed-field gel electrophoresis. About 18.5% of the library clones lacked inserts. The frequency of the BAC clones carrying chloroplast or mitochondrial DNA was 0.98% and 0.03%, respectively. The library represented approximately 4.9 haploid M. truncatula genomes. Hybridization of the BAC clone filters with a $C_{0}t-l$ DNA probe revealed that approximately 37% of the clones likely carried repetitive sequence-enriched DNA. An ordered array of pooled BAC DNA was screened by polymerase chain reactions using 13 sequence-characterized molecular markers that belonged to the eight linkage groups. Except for two markers, one to five positive BAC clones were obtained per marker. Accordingly, the sickle- and sunn-linked BAC clones identified herein will be useful for the isolation of these biotechnologically important genes. The new library will also provide clones that fill the gaps between preexisting BAC contigs, facilitating the physical mapping and genome sequencing of M. truncatula.

• Journal of Marine Bioscience and Biotechnology
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• v.2 no.1
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• pp.60-67
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• 2007

For the first time in Vietnam, morphological and molecular studies of a species belonging to Bacillariophyceae collected in Northern coast of Vietnam are presented. Observations with microscope showed that this species belong to genus: Pseudo-nitzschia and seem like P. pungens. Sequence data from the partial 18S small subunit ribosomal RNA gene (18S rDNA) and the internal transcribed spacer 1 - 5.8S - internal transcribed 2 have been used to determine clearly and generate a phylogenetic framework of the obtained sequences to previously reported sequences in GenBank. These results allowed us to highlight described species of Bacillariophyceae in Northern coast of Vietnam. Furthermore, accumulation of molecular study would be helpful for the identification of scientific name of harmful algal species and further taxonomic studies in Vietnam.

• Mycobiology
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• v.34 no.2
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• pp.45-55
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• 2006

Although Fursarium oxysporum causes diseases in economically important plant hosts, identification of F. oxysporum formae speciales has been difficult due to confusing phenotypic classification systems. To resolve these complexity, we evaluated genetic relationship of nine formae speciales of F. oxysporum with random amplified polymorphic DNA (RAPD), amplified fragment length polymorphism (AFLP), and translation elongation factor-l alpha ($EF-1{\alpha}$) gene. In addition, the correlation between mycotoxin content of fusaric acid and isolates based on molecular marker data was evaluated using the modified Mantel's test. According to these result, these fusaric acid-producing strains could not identify clearly, and independent of geographic locations and host specificities. However, in the identification of F. oxysporum formae speciales, especially, AFLP analysis showed a higher discriminatory power than that of a the RAPD and $EF-1{\alpha}$ analyses, all three techniques were able to detect genetic variability among F. oxysporum formae speciales in this study.

• Mycobiology
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• v.31 no.4
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• pp.235-247
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• 2003

In order to investigate biodiversity and establish identification system for Phytophthora spp. in Korea, a variety of band pattern was produced by using the URP(universal rice primer). The fingerprint patterns of Phytophthora spp. showed many common and variable fragments according to their isolates in distinct genotypes. In particular, P. drechsleri was classified into four distinct types(I to IV). P. drechsleri(KACC 40498 and KACC 40499) and P. cryptogea(KACC 40413) appeared to have almost equal bands despite their being different species. Ninety isolates of Phytophthora spp. were clustered into 13 groups based on UPGMA(unweighted pair group method with arithmetic means) analysis. These DNA fingerprinting data would be helpful for inter- and intra-species identification of Phytophthora species.

• Korean Journal of Ichthyology
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• v.25 no.2
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• pp.59-64
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• 2013

Two leptocephali (36.2~69.2 mm in total length) of the family Muraenidae were collected off Jeju Island, Korea. They were identified using 742 base pairs of mitochondrial DNA 12S rRNA sequences, and their morphological description were presented. The two leptocephali were identified to Gymnothorax minor based on genetic distance (d=0.000), being well consistent with that of G. minor adult collected off Tongyeong. Gymnothorax minor leptocephali are characterized by morphological combinations: total myomeres 137~139, predorsal myomeres 32, preanal myomeres 82~86, body depth in total length more than 10%.

• The Plant Pathology Journal
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• v.29 no.4
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• pp.357-363
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• 2013

The fungus Venturia nashicola is the causal agent of scab on Asian pears. For the rapid and reliable identification as well as sensitive detection of V. nashicola, a PCR-based technique was developed. DNA fingerprints of three closely related species, V. nashicola, V. pirina, and V. inaequalis, were obtained by random amplified polymorphic DNA (RAPD) analysis. Two RAPD markers specific to V. nashicola were identified by PCR, after which two pairs of sequence characterized amplified region (SCAR) primers were designed from the nucleotide sequences of the markers. The SCAR primer pairs, designated as D12F/D12R and E11F/E11R, amplified 535-bp and 525-bp DNA fragments, respectively, only from genomic DNA of V. nashicola. The specificity of the primer sets was tested on strains representing three species of Venturia and 20 fungal plant pathogens. The nested PCR primer pair specific to V. nashicola was developed based on the sequence of the species-specific 525-bp DNA fragment amplified by primer set E11F/E11R. The internal primer pair Na11F/Na11R amplified a 235-bp fragment from V. nashicola, but not from any other fungal species tested. The nested PCR assay was sensitive enough to detect the specific fragment in 50 fg of V. nashicola DNA.