• Journal of Life Science
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• v.27 no.7
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• pp.746-753
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• 2017

In order to rapidly identify four drums species, Larimichthys polyactis, L. crocea, Atrobucca nibe, and Pseudotolithus elongates, a highly efficient and quick method has been developed using multiplex polymerase chain reaction (PCR) with species-specific primers. Around 1.4 kbp of the mitochondrial COI gene sequences from the four drums species were aligned, and species-specific forward primers were designed, based on the single nucleotide polymorphism (SNP). The optimal conditions for PCR amplification were selected through cross-reactivity, using a gradient PCR method. The PCR results demonstrated species-specific amplification for each species at annealing temperatures between 50 and $62^{\circ}C$. Multiplex species-specific PCR (MSS-PCR) amplification reactions with four pairs of primers were performed for sixteen specimens of each species. MSS-PCR lead to a species-specific amplification of a 1,540 bp fragment in L. polyactis, 1,013 bp in A. nibe, 474 bp in L. crocea, and 182 bp in P. elongates, respectively. The four different sizes of each PCR product can be quickly and easily detected by single gel electrophoresis. The sensitivity of the MSS-PCR of the DNA was up to $0.1ng/{\mu}l$ as a starting concentration for the four different species tested. These results suggest that MSS-PCR, with species-specific primers based on SNP, can be a powerful tool in the rapid identification of the four drums species, L. polyactis, L. crocea, A. nibe, and P. elongates.

• Weed & Turfgrass Science
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• v.4 no.1
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• pp.26-34
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• 2015

Korean Poaceae includes approximately 80 species of the agricultural weeds. Precise species identification is the first step for more effective weed management in the agricultural fields. However, the identification of species in Poaceae is not easy without the assistance of taxonomists or identification experts although they are relatively easy to distinguish from the plants of the other family by the unique characteristics of caryopsis. Thus, DNA barcode was suggested as an alternative powerful technique for species identification by using short sections of DNA from a specific region of the genome. Two standard barcode markers of vascular plants, chloroplast rbcL and matK, and a supplementary nuclear ribosomal Internal Transcribed Spacer (ITS) region were used for barcode of major Korean Poaceae weeds, 403 individuals of 84 taxa. All the barcode markers revealed a good level of sequencing success with the lowest 73.7% for matK and the highest 88.8% for rbcL. The barcode sequences were deposited to the National Center for Biotechnology Information (NCBI) database for public use. Combined matK and ITS showed very high resolving power with 92.9%. Besides the identification of weeds for weed managment, the generated DNA barcode data could be used for many other applications such as rapid biodiversity assessment and conservation prioritization.

• Microbiology and Biotechnology Letters
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• v.31 no.2
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• pp.205-209
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• 2003

Recently, we introduced a new method for rapid screening of bacterial species- or subspecies-specific DNA probes, named “inverted dot blot hybridization screening method”. We then applied this method to develop species- or strain- specific DNA probes for Prevotella intermedia and Prevotella nigrescens. In those studies, among 96 candidate DNA probes which were screened by the new method, 5 probes were confirmed as being putatively strain-specific : 3 probes for P. nigrescens 9336 (ATCC 33563), one for each p. intermedia ATCC 25611 and one for P. nigrescens G8-9K-3 (ATCC 49046). In the present study, we evaluated by Southern blot analysis a DNA probe Pi29-L, one of the 96 candidate probes described above, whether it is specific for the strain ATCC 25611 off. intermedia. Our data show that the probe Pi29-L is potentially P. intermedia ATCC 25611-specific, which can be useful for the detection and identification of the strain, particularly in maintenance of the strain.

• Animal Systematics, Evolution and Diversity
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• v.37 no.1
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• pp.26-30
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• 2021

Botrylloides violaceus is native to the Northwest Pacific, including Korea. This species has many color variations in alive condition and a variety of zooid compound forms, and therefore difficult to identification in the field survey. This is the first report of COI DNA barcodes of B. violaceus from Korea. The intra-specific pairwise distance between Korean and UK populations had ranged from 1.4% to 2.6%. The inter-specific variations between B. violaceus and other Botrylloides species were 21.0-36.8%. The new DNA barcodes for Korean B. violaceus may be helpful in the identification of colonial ascidians, which is a difficult task when based on morphological identification.

• Parasites, Hosts and Diseases
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• v.55 no.6
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• pp.653-658
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• 2017

Hypoderma spp. larvae cause subcutaneous myiasis in several animal species. The objective of the present investigation was to identify and characterize morphologically and molecularly the larvae of Hypoderma spp. collected from cattle (Bos taurus taurus) and red deer (Cervus elaphus) in the district of Castelo Branco, Portugal. For this purpose, a total of 8 larvae were collected from cattle (n=2) and red deer (n=6). After morphological identification of Hypoderma spp. larvae, molecular characterization was based on PCR-RFLP and mitochondrial CO1 gene sequence analysis. All larvae were morphologically characterized as the third instar larvae (L3) of H. actaeon. Two restriction enzymes were used for molecular identification of the larvae. TaqI restriction enzyme was not able to cut H. actaeon. However, MboII restriction enzyme differentiated Hypoderma species showing 210 and 450 bp bands in H. actaeon. Furthermore, according to the alignment of the mt-CO1 gene sequences of Hypoderma species and to PCR-RFLP findings, all the identified Hypoderma larvae were confirmed as H. actaeon. This is the first report of identification of Hypoderma spp. (Diptera; Oestridae) from cattle and red deer in Portugal, based on morphological and molecular analyses.

• Parasites, Hosts and Diseases
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• v.54 no.6
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• pp.743-750
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• 2016

Mus musculus (Rodentia: Muridae) has generally been infected with a rodent hookworm Nippostrongylus brasiliensis. In this report, we present morphological and molecular identification of N. brasiliensis by light and scanning electron microscopy and PCR amplification of mitochondrial cytochrome c oxidase subunit 1 (cox1) gene and the protein sequences encoded by cox1 gene, respectively. Despite the use of N. brasiliensis in many biochemistry studies from India, their taxonomic identification was not fully understood, especially at the species level, and no molecular data is available in GenBank from India. Sequence analysis of cox1 gene in this study revealed that the present specimen showed close identity with the same species available in GenBank, confirming that the species is N. brasiliensis. This study represents the first record of molecular identification of N. brasiliensis from India and the protein structure to better understand the comparative phylogenetic characteristics.

• Proceedings of the Korean Institute of Information and Commucation Sciences Conference
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• 2013.05a
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• pp.1012-1014
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• 2013

DNA-DNA hybridization method with four oligonucleotide-specific probes was used simultaneously for differentiation and identification of four Mycobacterium species (Mycobacterium tuberculosis, M. avium, M. intracellulare, and M. kansasii). This DNA-DNA hybridization method with 4 oligonucleotide-specific probes, which targets in the rpoB region of 4 Mycobacteria species, respectively, was tested on 322 clinical isolates. Using DNA-DNA hybridization method, we detected M. tuberculosis (282 strains), M. avim (7 strains), M. intracellulare (9 strains), and M. kansasii (3 strain) from 322 clinical isolates. This result was compared with conventional biochemical test and rpoB DNA sequence analysis of this clinical isolates. We confirmed identification of Mycobacterium tuberculosis, M. avium, M. intracellulare, and M. kansasii with high sensitivity (100 %) and specificity (100 %). This DNA-DNA hybridization method could be performed within 4 hours at least. Therefore, we suggest that DNA- DNA hybridization method using 4 rpoB DNA probes of Mycobacteria could be used for accurate, rapid, convenient detection and identification of Mycobacterium tuberculosis, M. avium, M. intracellulare, and M. kansasii in clinical samples.

• Korean Journal Plant Pathology
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• v.14 no.6
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• pp.603-605
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• 1998

Soil and root samples were collected form the rhizoshpere of 11 different medicinal plants to determine the incidence, density and identification of root-knot nematode species associated with medicinal herbs. About 55% of medicinal herbs examined was found to be infested with root-knot nematodes. As a result of infection casued by three root-knot nematodes, M. hapla recorded 43.3% in medicinal herba whereas M. incognita and M. arenaria showed 7.9% and 3.7%, repectively. Forsythia koreana, Hemerocalis fulva, Hibuscus mutabilis and Petasites japonicus were the most severely infested herbs whereas Acanthopanax sessilflorus was least infested. Population of the second stage younger plants. Meloidogyne hapla, M. incognita and M. arenaria were the species associated with the medicinal herbs. The most abundant nematode observed in medicinal herbs was M. hapla and followed by M. incognita and M. arenaria. M. arenaria was observed firstly on Ficus carica, one of medicinal plant.

• Parasites, Hosts and Diseases
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• v.60 no.4
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• pp.273-279
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• 2022

Laelapinae mites are involved in transmission of microbial diseases between wildlife and humans, with an impact on public health. In this study, 5 mite members in the subfamily Laelapinae (laelapin mites; LM) were morphologically identified by light microscopy, and the phylogenetic relationship of LM was analyzed in combination with the sequence information of part of the LM cytochrome oxidase subunit I (cox1) gene. The morphological identification revealed that 5 mites belonged to the genera Laelaps and Haemolaelaps, respectively. Sequence analysis showed that the ratio of nonsynonymous mutation rate to synonymous mutation rate of LM was less than 1, indicating that the LM cox1 gene had undergone purifying selection. Phylogenetic analysis showed that the Laelapinae is a monophyletic group. The genera Haemolaelaps and Hyperlaelaps did not separated into distinct clades but clustered together with species of the genus Laelaps. Our morphological and molecular analyses to describe the phylogenetic relationships among different genera and species of Laelapinae provide a reference for the improvement and revision of the LM taxonomy system.

• The Korea Journal of Herbology
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• v.39 no.2
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• pp.1-9
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• 2024

Objectives : Paeng-hwal is described as an insect herbal medicine used for digestive diseases in the Dong-ui-bo-gam. The origin of this herbal medicine is limited to several small crabs, such as Helice tridens. These crab species cohabitat in the same environment and share similar morphological characteristics, making it very difficult to distinguish and collect the individual species for use in dietary supplements or herbal medicines. This study was conducted to develop a genetic identification tool for discriminating among these closely related small crab species. Methods : CO1 DNA barcode regions of 15 samples from 6 species of small crabs were analyzed to obtain the individual sequences. To identify the correct species, comparative analyses were carried out using the database of the NCBI GenBank and the NIBR. SCAR primers were designed to develop simple and rapid assay methods using inter-species specific sequences. Optimal SCAR assay conditions were established through gradient PCR, and the limit of detection (LOD) was determined. Results : Six species of small crabs (Helicana tridens, Macrophthalmus abbreviatus, Helicana tientsinensis, Helicana wuana, Chiromantes dehaani, and Hemigrapsus penicillatus), which are distributed as Paeng-hwal, were identified through CO1 sequences analysis. We also developed SCAR markers to distinguish between six small crabs at the species level. Furthermore, we established the optimal PCR assay methods and the LOD of each individual species. Conclusions : The rapid and simple SCAR-PCR assay methods were developed to identify the species and control the quality of herbal medicines for Paeng-hwal based on the genetic analyses of CO1 DNA barcodes.