• Food Science and Biotechnology
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• v.18 no.3
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• pp.694-699
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• 2009

A multiplex polymerase chain reaction (PCR) method was developed for the detection of 4 events of genetically modified (GM) soybean. The event-specific primers were designed from 4 events of GM soybean (RRS, A2704-12, DP356043-5, and MON89788). The lectin was used as an endogenous reference gene of soybean in the PCR detection. The primer pair YjLec-4-F/R producing 100 bp amplicon was used to amplify the lectin gene and no amplified product was observed in any of the 9 different plants used as templates. This multiplex PCR method allowed for the detection of event-specific targets in a genomic DNA mixture of up to 1% GM soybean mixture containing RRS, A2704-12, DP356043-5, and MON89788. In this study, 20 soybean products obtained from commercial food markets were analyzed by the multiplex PCR. As a result, 6 samples contained RRS. These results indicate that this multiplex PCR method could be a useful tool for monitoring GM soybean.

• Journal of Microbiology and Biotechnology
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• v.29 no.10
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• pp.1543-1552
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• 2019

Salmonella is a common zoonotic and foodborne pathogen that causes high morbidity and mortality in developing countries. In this study, we established and validated a polymerase spiral reaction (PSR) assay which targeted the conserved invasion gene (invA) of Salmonella by SYBR Green I indicator methods. Subsequently, assays for determination of the optimal conditions for optimal specificity and sensitivity of PSR were performed. We performed comprehensive evaluations using loop-mediated isothermal amplification (LAMP) and real-time PCR. A total number of 532 samples of daily food were analyzed by PSR. Twenty-seven bacterial strains were tested in the specificity assay, from which positive results were obtained only for 14-Salmonella strains. However, none of the 13 non-Salmonella strains was amplified. Similarly with LAMP and real-time PCR, the detection limit of the PSR assay was 50 CFU/ml. The PSR method was also successfully applied to evaluate the contamination with Salmonella in 532 samples of daily food, corroborating traditional culture method data. The novel PSR method is simple, sensitive, and rapid and provides new insights into the prevention and detection of foodborne diseases.

• Journal of Microbiology and Biotechnology
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• v.34 no.3
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• pp.681-688
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• 2024

The accurate and rapid detection of methicillin-resistance of Staphylococcus aureus (SA) holds significant clinical importance. However, the methicillin-resistance detection strategies commonly require complicated cell lysis and gene extraction. Herein, we devised a novel colorimetric approach for the sensitive and accurate identification of methicillin-resistance of SA by combining allosteric probe-based target recognition with self-primer elongation-based target recycling. The PBP2a aptamer in the allosteric probe successfully identified the target MRSA, leading to the initiation of self-primer elongation based-cascade signal amplification. The peroxidase-like hemin/G-quadruplex undergo an isothermal autonomous process that effectively catalyzes the oxidation of ABTS2- and produces a distinct blue color, enabling the visual identification of MRSA at low concentrations. The method offers a shorter duration for bacteria cultivation compared to traditional susceptibility testing methods, as well as simplified manual procedures for gene analysis. The overall amplification time for this test is 60 min, and it has a detection limit of 3 CFU/ml. In addition, the approach has exceptional selectivity and reproducibility, demonstrating commendable performance when tested with real samples. Due to its advantages, this colorimetric assay exhibits considerable potential for integration into a sensor kit, thereby offering a viable and convenient alternative for the prompt and on-site detection of MRSA in patients with skin and soft tissue infections.

• The Zoological Society Korea : Newsletter
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• v.12 no.2
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• pp.126.2-126
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• 1995

No Abstract, See Full Text

• Proceedings of the Zoological Society Korea Conference
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• 1995.10b
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• pp.302.2-302
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• 1995

No Abstract, See Full Text

• Proceedings of the Zoological Society Korea Conference
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• 1995.10b
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• pp.345.2-345
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• 1995

No Abstract, See Full Text

• Journal of the Korean Society of Food Science and Nutrition
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• v.41 no.9
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• pp.1326-1330
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• 2012

This study was conducted to develop a loop-mediated isothermal amplification (LAMP) method for the detection of Clostridium difficile. The tested target gene was 16S ribosomal RNA. Five different LAMP primer sets were designed, and LAMP was performed. All primer sets targeting the 16S rRNA gene (BIP, FIP, B3, F3, LF, PF) were determined as positive in tcdA-positive, tcdB-postive ($A^+B^+$) and tcdA-negative, tcdB-negative ($A^-B^-$) Clostridium difficile strains. As the LAMP reaction took less than 80 min and did not require expensive machine such as thermocycler, it can be used as a rapid and simple detection method for foodborne pathogens.

• Korean Journal of Veterinary Research
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• v.39 no.2
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• pp.343-352
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• 1999

Vancomycin resistant enterococci (VRE) have emerged as an important nosocomial pathogen. Since 1989 the Center for Disease Control, United States, has reported a rapid increase in the incidence of enterococcal bacteremia and endocarditis infection by VRE. It was suggested that the use of avoparcin was associated with the appearance of VRE in animal husbandry. To date, several detection methods have been used based on conventional methods of culture and gene detection. However, these methods have some limitations such as time-consuming, laborious and additional differential needs. Therefore, In this study a multiplex PCR method was established to detect and differentiate resistance types of enterococci which specifically amplify the four van genes encoding vancomycin resistance elements. Using the method, we investigated the incidence rates and types of VRE from farms using or not using avoparcin. A total of 1091 animal fecal samples were collected from 70 pig and 32 poultry farms. A total of 425 of enterococci were isolated from samples. Of the 425 isolates, 11 of the them showed a pattern of high-level vancomycin resistance (MIC : $64{\sim}256{\mu}g/ml$) which was associated with the presence of the vanA or vanB gene. Fifty-seven isolates showed a pattern of low-level vancomycin resistance (MIC : $3{\sim}8{\mu}g/ml$) associated with the vanC-1 or vanC-2 gene. Interestingly, all isolates with high-level vancomycin resistance were from farms that have never used avoparcin. Moreover, the high-level VRE isolation rate in Korea (2.58%) was much lower than that of other countries (50% in England, 7% in Belgium) where avoparcin have been used. In conclusion, the multiplex PCR method established in this study could be applied for detection of VRE.

• Journal of Microbiology and Biotechnology
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• v.21 no.7
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• pp.719-727
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• 2011

Botulism is a neuroparalytic disease caused by Clostridium botulinum, which produces seven (A-G) antigenically diverse neurotoxins (BoNTs). BoNTs are the most poisonous substances known to humans, with a median lethal dose ($LD_{50}$) of approximately 1 ng/kg of body weight. Owing to their extreme potency and lethality, they have the potential to be used as a bioterrorism agent. The mouse bioassay is the gold standard for the detection of botulinum neurotoxins; however, it requires at least 3-4 days for completion. Attempts have been made to develop an ELISA-based detection system, which is potentially an easier and more rapid method of botulinum neurotoxin detection. The present study was designed using a synthetic gene approach. The synthetic gene encoding the catalytic domain of BoNT serotype B from amino acids 1-450 was constructed with PCR overlapping primers (BoNT/B LC), cloned in a pQE30 UA vector, and expressed in an E. coli M15 host system. Recombinant protein production was optimized at 0.5 mM IPTG final concentration, 4 h post induction, resulting in a maximum yield of recombinant proteins. The immunogenic nature of the recombinant BoNT/B LC protein was evaluated by ELISA. Antibodies were raised in BALB/c mice using various adjuvants. A significant rise in antibody titer (p<0.05) was observed in the Alum group, followed by the Titermax Classic group, Freund's adjuvant, and the Titermax Gold group. These developed high-titer antibodies may prove useful for the detection of botulinum neurotoxins in food and clinical samples.

• 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.