• 대한임상검사과학회지
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• 제38권3호
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• pp.173-178
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• 2006

The production of extended-spectrum ${\beta}$-lactamases ($ESBL_S$) is the main mechanism of bacterial resistance to third-generation cephalosporins and monobactams, whose prevalence varies depending on the different geographical areas. In the last years it has increased notably to the point of being considered a health problem of great importance. The characterization of the ESBLs producing Klebsiella penumoniae strains present in clinical isolates is time-consuming. I describe here the development of a new system, which consists of a multiplex PCR. I found 51 K. pneumoniae strains to be presumptive strains ESBLs producers by clinical and laboratory standards institute (CLSI) guidelines. The double disc synergy test showed 47 positive K. pneumoniae, which were K. pneumoniae isolates. All ESBLs producing K. pneumoniae strains were resistant to antibiotic amikacin, gentamicin and ciprofloxacin. By multiplex PCR analysis, $bla_{TEM}$ gene in 17 strains 44 $bla_{SHV}$ genes and $bla_{CTX}$ genes in 33 strains were identified. In this study, the multiplex polymerase chain reaction (PCR) assay was a good method to detect and differentiate ESBLs producing K. penumoniae strains in clinical isolates.

• Clinical and Experimental Pediatrics
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• 제55권11호
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• pp.424-429
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• 2012

Purpose: Methods for quick and reliable detection of Streptococcus pneumoniae are needed for the diagnosis of pneumococcal disease and vaccine studies. This study aimed to show that sequential multiplex polymerase chain reaction (PCR) is more efficient than conventional culture in achieving S. pneumoniae -positive results. Methods: Nasopharyngeal (NP) secretions were obtained from 842 pediatric patients admitted with lower respiratory infections at Severance Children's Hospital in Korea between March 2009 and June 2010. For identification and serotype determination of pneumococci from the NP secretions, the secretions were evaluated via multiplex PCR technique with 35 serotype-specific primers arranged in 8 multiplex PCR sets and conventional bacteriological culture technique. Results: Among the results for 793 samples that underwent both bacterial culture and PCR analysis for pneumococcal detection, 153 (19.3%) results obtained by PCR and 81 (10.2%) results obtained by conventional culture technique were positive for S. pneumoniae. The predominant serotypes observed, in order of decreasing frequency, were 19A (23%), 6A/B (16%), 19F (11%), 15B/C (5%), 15A (5%), and 11A (4%); further, 26% of the isolates were non-typeable. Conclusion: As opposed to conventional bacteriological tests, PCR analysis can accurately and rapidly identify pneumococcal serotypes.

• Journal of Microbiology
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• 제39권3호
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• pp.226-228
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• 2001

Human lymphotropic herpesvirus is known to be a major pathogen associated with various diseases in bone marrow transplantation (BMT) recipients. A multiplex nested-polymerase chain reaction (PCR) method was developed for the simultaneous detection of human lymphotropic herpesviruses, including Ebstein-Barr virus (EBV), cytomegalovirus (CMV), and human herpesvirus 6 variants A and B (HHV6-A, HHV6-B). To demonstrate the usefulness of multiplex PCR for the analysis of clinical samples, peripheral blood mononuclear cells and serum from BMT recipients were analysed. The results skewed that a clear detection could be made between EBV, HCMV and HHV-6. This multiplex PCR assay is an efficient and cost-effective approach to the analysis of large numbers of samples to determine the epidemiological importance of EBV HCMV and HHV-6.

• Journal of Microbiology
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• 제40권2호
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• pp.146-150
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• 2002

A multiplex polymerase chain reaction (PCR) assay was developed for the identification of two Candida species-albicans and dubliniensis. Three sets of primers were selected from different genomic sequences to specifically amplify a 516 bp fragment within the tops gene, specific for several species of the genus Candida (CCL primers); a 239 bp fragment within the $\alpha$INT1 gene, specific for Candida albicans (CAL primers); and a 175 bp fragment within the ALSD1 gene, specific for Candida dubliniensis (CDL primers). Using the primers in conjunction (multiplex PCR), we were able to detect both C. albicans and C. dubliniensis and to differentiate between them. The detection limit of the PCR assay was approximately 10 cells per milliliter of saline. Thus, this multiplex PCR assay can be applied for differentiation of C. albicans and C. dubliniensis from clinical specimens.

• 한국약용작물학회지
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• 제26권1호
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• pp.26-31
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• 2018

Background: Angelica gigas, A. sinensis, and A. acutiloba are commercially important in the herbal medicine market, and among them, A. gigas has the highest economic value and price. However, their similar morphological traits are often used for fraud. Despite their importance in herbal medicine, recognition of the differences between Angelica species is currently inadequate. Methods and Results: A multiplex polymerase chain reaction (PCR) method was developed for direct detection and identification of A. gigas, A. sinensis, and A. acutiloba. The gene for the distinction of species was targeted at ITS in the nucleus and trnC-petN gene in chloroplasts. The optimized multiplex PCR in the present study utilized each Angelica species-specific primer pairs. Each primer pair yielded products of 229 base pairs (bp) for A. gigas, 53 bp for A. sinensis, 170 bp for A. acutiloba. Additionally non-specific PCR products were not detected in similar species by species-specific primers. Conclusions: In the present study, a multiplex-PCR assay, successfully assessed the authenticity of Angelica species (A. gigas, A. sinensis, and A. acutiloba). and whole genome amplification (WGA) was performed after DNA extraction to identify, the species in the product. The detection method of raw materials developed in the present study could be applied to herbal medicine and health functional food management.

• 식물병연구
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• 제24권4호
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• pp.348-352
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• 2018

A multiplex reverse transcription-polymerase chain reaction (mRT-PCR) assay was developed for the detection of Clover yellow vein virus (ClYVV), Peanut mottle virus (PeMoV), and Tomato spotted wilt virus (TSWV), which were recently reported to infect soybean and azuki bean in Korea. Species-specific primer sets were designed for the detection of each virus, and their specificity and sensitivity were tested using mixed primer sets. From among the designed primer sets, two combinations were selected and further evaluated to estimate the detection limits of uniplex, duplex, and multiplex RT-PCR. The multiplex RT-PCR assay could be a useful tool for the field survey of plant viruses and the rapid detection of ClYVV, PeMoV, and TSWV in leguminous plants.

• 생명과학회지
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• 제21권7호
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• pp.947-952
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• 2011

현재까지 다수의 역학연구를 통해 피부에 발생한 보통사마귀에서 제 1, 2, 3, 4, 7, 10, 27, 57 및 65형의 인유두종바이러스가 검출되었다. 그러나 기존의 중합효소연쇄반응(conventional polymerase chain reaction, PCR)을 이용하는 경우 절차가 복잡하여 시간이 오래 걸리는 단점이 있었다. 이번 연구를 통해 저자들은 보통사마귀에서 가장 흔히 검출되는 6가지 유전자형의 인유두종바이러스를 한번에 확인 가능한 간편한 muliplex PCR의 개발을 목표로 하였다. 인유두종바이러스의 염기서열분석을 통해, L1에서 E6, 그리고 E2에서 L2 사이의 유전자간영역(intergenic region)으로 부터 6쌍의 primer를 고안하였으며, L1 유전자서열 분석을 통해 multiplex PCR의 특이성을 확인하였다. 총 129개의 조직표본 중 109개에서 제 1, 2, 3, 4, 27, 57형의 인유두종바이러스를 확인하였다. 이번 연구의 primer를 이용한 인유두종바이러스 검출의 민감도와 특이도는 각각 85%와 99.5%였다. 이러한 primer 세트로 인유두종바이러스가 검출되지 않은 20개의 조직표본의 경우, 또 다른 HPV primer를 사용한 추가적인 multiplex PCR을 시행하여 7개 표본에서 제 7형 및 65형의 인유두종바이러스가 검출되었다. 이상의 결과는 본 연구를 통해 새롭게 고안된 multiplex PCR 기법을 통해 보통사마귀에서의 인유두종바이러스를 보다 정확하고 빠르게 검출할 수 있다는 것을 보여 준다.

• Food Science and Biotechnology
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• 제17권3호
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• pp.569-572
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• 2008

A multiplex polymerase chain reaction (PCR) method was developed to simultaneously detect 3 events of genetically modified (GM) maize. The event-specific primers were used to discriminate the following 3 events of GM maize (DAS-59122-7, TC6275, and MIR604) using multiplex PCR method. The zein gene was used as an endogenous maize reference gene in the multiplex PCR detection. The primer pair Zein-FIR producing a 99 bp amplicon was used to amplify the zein gene. The primer JI-Das-F1/R1 for DAS-59122-7, JI-TC6275-F3/R3 for TC6275, and JI-MIR F1/R1 for MIR604 yielded an amplicon of 130, 162, and 197 bp, respectively. The detection limit of multiplex PCR was 1% for DAS-59122-7, TC6275, and MIR604 for one reaction.

• 대한수의학회지
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• 제43권1호
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• pp.103-112
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• 2003

A multiplex PCR assay, which allows simultaneous detection of vancomycin resistant genotypes and Enterococcus species-specific genes, was developed. Vancomycin resistant enterococci (VRE) from chickens and humans could be detected for vanA, vanB, vanC-1, vanC-2, $ddl_{E.faecium}$ and $ddl_{E.faecalis}$ by multiplex PCR. Eight isolates of VRE from humans (n=11) had $ddl_{E.faecium}$ and vanA, and 3 isolates of the VRE had $ddl_{E.faecium}$ and vanB. One isolate of VRE from chickens (n=6) had $ddl_{E.faecium}$ and vanA, and 5 isolates of the VRE had only vanA. E. faecium, E. faecalis, E. gallinarum and E. casseliflavus were also confirmed for the species-specific gene by multiplex PCR. This multiplex PCR could detect E. faecium, E. faecalis, E. gallinarum, E. casseliflavus, vanA, vanB, vanC-1 and vanC-2, simultaneously. The PCR assay established in the present study can be an alternative to time-consuming biochemical tests and antibiotic susceptibility tests of Enterococcus spp.

• Proceedings of the National Institute of Ecology of the Republic of Korea
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• 제1권1호
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• pp.83-89
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• 2020

Methods for detecting the presence of genetically modified (GM) crops are evolving to comply with legislation and to enhance monitoring by biotechnology companies and regulators. In order to cover a broad range of detection methods for a new GM crop, conventional multiplex PCR methods are required. Based on the genetic information on three GM alfalfa varieties (J101, J163, and KK179), which were recently approved in South Korea, we developed a fast, reliable, and highly specific multiplex polymerase chain reaction (PCR) method with basic PCR equipment and inexpensive reagents. To validate and verify the newly developed multiplex PCR method, we applied a limit of detection assay and random reference material analysis. We also monitored the unintentional environmental release of GM alfalfa in South Korea by performing the multiplex PCR analysis with 91 feral alfalfa specimens collected from 2000 to 2018. Our methodology is a sensitive, simple, quick, and inexpensive tool for detecting and identifying three GM alfalfa varieties.