• Journal of Microbiology
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• v.39 no.2
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• pp.126-132
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• 2001

For rapid and secure differentiation of P. infestans from other Phytophthora species, two fragments obtained from randomly amplified polymorphic DNA (RAPD) profiles were selected as markers. Also, primers for in polymerase chain reaction (PCR) to detect P. infestans specifically were developed by analyzing the sequences of ITSII regions in rDNA of Phytophthora species. The primers, PISP-1 and ITS3 amplified a single. Fragment 450 bp of about in P. infestans, but not in other fungal or bacterial isolates. Annealing temperatures and template DNA quantities were varied for the optimization of PCR conditions. From the result of the PCR detection study, species-specific primers were selected under annealing temperatures ranging from 55$^{\circ}C$ to 61$^{\circ}C$, and template DNA levels ranging from 10 pg to 100 ng.

• The Korean Journal of Mycology
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• v.44 no.2
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• pp.103-107
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• 2016

To establish a rapid and accurate detection of Phytophthora pinifolia, which is a quarantine pathogenic fungus in Korea, a species-specific primer was developed based on the ras-related protein (Ypt1) gene. Species-specific primer based on the DNA sequences of Ypt1 gene amplified 193 bp polymerase chain reaction (PCR) product for P. pinifolia. The primer pair yielded the predicted PCR product size exactly in testing with target pathogen DNAs, but not from the other 10 species of Phytophthora and 14 species of other phytopathogenic fungi. The primer pair also showed only the species-specific amplification curve on realtime PCR on target pathogen DNA. The detection sensitivity of real time PCR using species-specific primer pair was 10 to 100 times higher than conventional PCR, with 1 to $10pg/{\mu}L$.

• Journal of Dairy Science and Biotechnology
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• v.33 no.4
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• pp.253-262
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• 2015

The authentication and implications of misleading labeling in milk and dairy products is important to protect against cheating consumers from adulteration and to alert sensitive consumers to any undeclared potential allergens. This need to support milk and dairy products labeling has led to the development of specific analytical techniques for the analysis of milk and dairy products ingredients. Recently, several methods based on polymerase chain reaction (PCR), including restriction fragment length polymorphism (PCR-RFLP), multiplex PCR, species-specific PCR, and real-time PCR, have been proposed as useful means for identifying species of origin in milk and dairy products, as well as quantifying and detecting any adulteration. These methods have particular advantages owing to their high specificity and sensitivity, as well as rapid processing time. In this review, we provide an updated and extensive overview of the PCR-based methods used for milk and dairy products authentication with a particular focus on the application of PCR methods to detect adulteration.

• Proceedings of the National Institute of Ecology of the Republic of Korea
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• v.3 no.4
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• pp.212-220
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• 2022

DNA markers have been studied and used intensively to identify plant species based on molecular approaches. The genus Medicago belongs to the family Fabaceae and contains 87 species distributed from the Mediterranean to central Asia. Five species of Medicago are known to be distributed in South Korea; however, their morphological characteristics alone cannot distinguish the species. In this study, we analyzed the phylogenetic relationships using collected five species of Medicago from South Korea and 44 taxa nucleotide information from NCBI. The constructed phylogenetic tree using gibberellin 3-oxidase 1 and tRNA^Lys (UUU) to maturase K gene sequences showed the monophyly of the genus Medicago, with five species each forming a single clade. These results suggest that there are five species of Medicago distributed in South Korea. In addition, we designed polymerase chain reaction primers for species-specific detection of Medicago by comparing the plastid sequences. The accuracy of the designed primer pairs was confirmed for each Medicago species. The findings of this study provide efficient and novel species identification methods for Medicago, which will assist in the identification of wild plants for the management of alien species and living modified organisms.

• Food Science and Biotechnology
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• v.17 no.6
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• pp.1240-1245
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• 2008

Kimbab is the most popular ready-to-eat (RTE) food in Korea. A rapid detection method based on multiplex PCR technique was developed for detection of major food-borne pathogens like Salmonella spp., Shigella spp., Bacillus cereus, Listeria monocytongenes, and Staphylococcus aureus. Specific bands were obtained as 108 bp (Sau, S. aureus), 284 bp (Sal, S. enterica, S. enteritids, and S. typhmurium), 404 bp (Lmo, L. monocytogenes), 475 bp (Bce, B. cereus), and 600 bp (Shi, S. flexineri and S. sonnei). Visible cell numbers varied from 4.14-5.03, 3.61-4.47, and 4.10-5.11 log CFU/g in randomly collected June, July, and August samples, respectively. Among the 30 kimbab samples obtained 83.3% samples were contaminated and 16.7% samples were free from contamination. The highest rate of contamination was with S. aureus (56.7%) followed by B. cereus (43.3%), Salmonella spp. (36.7%), Shigella spp. (13.3%), and L. monocytogenes (6.7%). The identification of the pathogenic species could be faster using one polymerase chain reaction (PCR) and the ability to test for food-borne pathogenic species in kimbab will save time and increase the ability to assure its quality.

• Korean Journal of Veterinary Service
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• v.38 no.2
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• pp.101-106
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• 2015

Murine mycoplasmosis, caused by Mycoplasma (M.) pulmonis, is a prominent disease in rodent animals. The aim of this study was to develop a sensitive and specific PCR assay to detect M. pulmonis in animals and to assess the suitability of this assay for the detection of mycoplasmal infection in rats experimentally infected with M. pulmonis. A new PCR assay using the M. pulmonis-specific primer pairs MPul-F and MPul-R was developed. The primers and probe for the assay were designed from regions in the 16S rRNA gene that are unique to M. pulmonis. The novel PCR assay was very specific and sensitive for M. pulmonis, detecting the equivalent of 5 pg of target template DNA. It detected only M. pulmonis and no other Mycoplasma species or other bacterial species. The newly developed PCR assay also effectively detected M. pulmonis infection in rats. These results suggest that this PCR assay using M. pulmonis-specific primer pairs of MPul-F and MPul-R will be useful and effective for monitoring M. pulmonis infection in animals.

• The Plant Pathology Journal
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• v.38 no.6
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• pp.629-636
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• 2022

Colletotrichum species is known as the major causal pathogen of red pepper anthracnose in Korea and various groups of fungicides are registered for the management of the disease. However, the consistent use of fungicides has resulted in the development of resistance in many red pepper-growing areas of Korea. Effective management of the occurrence of fungicide resistance depends on constant monitoring and early detection. Thus, in this study, various methods such as agar dilution method (ADM), gene sequencing, allele-specific polymerase chain reaction (PCR), and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) were applied for the detection of benzimidazole resistance among 24 isolates of Colletotrichum acutatum s. lat. and Colletotrichum gloeosporioides s. lat. The result of the ADM showed that C. gloeosporioides s. lat. was classified into sensitive and resistant isolates to benomyl while C. acutatum s. lat. was insensitive at ≥1 ㎍/ml of benomyl. The sequence analysis of the β-tubulin gene showed the presence of a single nucleotide mutation at the 198th amino acid position of five isolates (16CACY14, 16CAYY19, 15HN5, 15KJ1, and 16CAYY7) of C. gloeosporioides s. lat. Allele-specific PCR and PCR-RFLP were used to detect point mutation at 198th amino acid position and this was done within a day unlike ADM which usually takes more than one week and thus saving time and resources that are essential in the fungicide resistance management in the field. Therefore, the molecular techniques established in this study can warrant early detection of benzimidazole fungicide resistance for the adoption of management strategies that can prevent yield losses among farmers.

• ALGAE
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• v.39 no.1
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• pp.51-56
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• 2024

The use of droplet digital polymerase chain reaction (ddPCR) has greatly improved the quantification of harmful protists, outperforming traditional methods like quantitative PCR. Notably, ddPCR provides enhanced consistency and reproducibility at it resists PCR inhibitors commonly found in environmental DNA samples. This study aimed to determine the most effective filter material for ddPCR protocols by assessing the reproducibility of species-specific gene copy numbers and filtration time across six filter types: cellulose acetate (CA), mixed cellulose ester (MCE), nylon (NY), polycarbonate (PC), polyethersulfone (PES), and polyvinylidene fluoride (PVDF). The study used two species of Chattonella marina complexes as a case study. Filtration rates were slower for NY, PC, and PVDF filters. Moreover, MCE, NY, PES, and PVDF yielded lower DNA amounts than other filters. Importantly, the CA filter exhibited the lowest variance (38-39%) and the highest determination coefficients (R² = 0.92-0.96), indicating superior performance. These findings suggest that the CA filter is the most suitable for ddPCR quantification of marine protists, offering quick filtration and reliable reproducibility.

• Journal of Microbiology and Biotechnology
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• v.22 no.8
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• pp.1113-1117
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• 2012

Shigella sonnei is a causal agent of fever, nausea, stomach cramps, vomiting, and diarrheal disease. The present study describes a quantitative polymerase chain reaction (qPCR) assay for the specific detection of S. sonnei using a primer pair based on the methylase gene for the amplification of a 325 bp DNA fragment. The qPCR primer set for the accurate diagnosis of Shigella sonnei was developed from publically available genome sequences. This quantitative PCR-based method will potentially simplify and facilitate the diagnosis of this pathogen and guide disease management.

• Journal of Dairy Science and Biotechnology
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• v.31 no.2
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• pp.127-131
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• 2013

The aim of this study was to investigate adulteration of caprine milk products by bovine milk using biomolecular techniques with bovine-specific primers for the mitochondrial cytochrome b gene. Polymerase chain reaction (PCR) and real-time PCR assays were applied to caprine milk products including infant formula, city milk, and fermented milk. The results indicated that six out of the eight caprine infant formula products tested contained bovine milk components. In addition, two of the three tested caprine city milk products and two caprine fermented milk products were shown to be adulterated with bovine milk. Conventional PCR results corroborated with results obtained by real-time PCR. This study demonstrates that DNA-based species identification procedures would be useful and applicable in routine examinations of the dairy industry to ensure the quality and safety of dairy foods.