• Food Science and Preservation
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• v.16 no.6
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• pp.965-970
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• 2009

PCR technology has been widely used to detect and quantify microbial pathogens in foodstuffs, because the technique is rapid, sensitive, and selective. In this study, detection of contaminating pathogenic bacteria on shells of chicken eggs was performed using both a commercial multiplex polymerase chain reaction (PCR) kit and a viable count method employing a selective medium. The PCR kit was capable of detecting Campylobacter jejuni, Escherichia coli O157:H7, Staphylococcus aureus, Bacillus cereus, Vibrio parahaemolyticus, Listeria monocytogenes, Yersinia enterocolitica, Salmonella species, and Shigella species. Using the PCR method, five bacterial species were detected from 30 samples (33.3%) of 90 batches of eggs commercially available in a market. PCR products from B. cereus, S. aureus, L. monocytogenes, Y. enterocolitica, and E. coli O157:H7 were detected, and the numbers and frequencies of positive samples were 17 (18.8%), 12 (13.3%), 15 (16.6%), 16 (17.7%),and 4 (4.4%), respectively. None of any Salmonella species, C. jejuni, V. parahaemolyticus, or Shigella species was detected in this study. The results of PCR testing were confirmed using a typical viable count method employing a selective medium. We suggest that the multiplex polymerase chain reaction (mPCR) assay is a rapid and reliable method for detection of pathogenic bacteria contaminating eggs.

• Korean Journal of Clinical Laboratory Science
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• v.39 no.3
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• pp.161-166
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• 2007

To develop a rapid and reliable single-tube-based PCR technique for detection simultaneously the quinolone resistance qnrA, qnrB and qnrS genes. After multiple alignment, primers were designed to detect known qnr variants. I was used for A total of 43 extented-spectrum ${\beta}$-lactamases (ESBLs) producing Escherichia coli and Klebsiella pneumoniae isolated from university hospital were tested for screening, as with qnr genes. In optimized conditions, all positive controls confirmed the specificity of the PCR primers. Out of 43 isolates, qnrA genes were detected 19 (44.2%), qnrB genes 5 (11.7%), qnrS genes 15 (34.9%) and 8 (18.6%) isolates were not detected. I report here a fast and reliable technique for rapid screening of qnr positive strains to be used for epidemiological surveys.

• Proceedings of the Microbiological Society of Korea Conference
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• 2005.05a
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• pp.203.2-203
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• 2005

• Korean Journal of Veterinary Service
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• v.37 no.4
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• pp.247-252
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• 2014

The economic impact of swine mycoplasma infection is high. An accurate diagnosis is often difficult and time consuming. We report the development and validation of an effective multiplex polymerase chain reaction (PCR) assay that detects Mycoplasma (M.) hyopneumoniae and M. hyorhinis. The multi detection of M. hyopneumoniae and M. hyorhinis primer set were employed to detect mycoplasma species and typing of the species was performed on the basis of sequence analysis of the PCR product. The target nucleic acid fragments were specifically amplified by M. hyopneumoniae and M. hyorhinis PCR with 16S ribosomal DNA primers. Single and mixed Mycoplasma species DNA templates were used to evaluate the specificity of the multiplex assay. The corresponding specific DNA products were amplified for each pathogen. The multiplex PCR assay provides a novel tool for simultaneous detection and differentiation of M. hyopneumoniae and M. hyorhinis.

• The Plant Pathology Journal
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• v.29 no.3
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• pp.338-343
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• 2013

A detection system based on a multiplex reverse transcription (RT) polymerase chain reaction (PCR) was developed to simultaneously identify multiple viruses in the lily plant. The most common viruses infecting lily plants are the cucumber mosaic virus (CMV), lily mottle virus (LMoV), lily symptomless virus (LSV). Leaf samples were collected at lily-cultivation facilities located in the Kangwon province of Korea and used to evaluate the detection system. Simplex and multiplex RT-PCR were performed using virus-specific primers to detect single- or mixed viral infections in lily plants. Our results demonstrate the selective detection of 3 different viruses (CMV, LMoV and LSV) by using specific primers as well as the potential of simultaneously detecting 2 or 3 different viruses in lily plants with mixed infections. Three sets of primers for each target virus, and one set of internal control primers were used to evaluate the detection system for efficiency, reliability, and reproducibility.

• Food Science of Animal Resources
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• v.31 no.5
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• pp.658-662
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• 2011

In this study, 543 samples of press hams, sausages, processed ground meat and processed cheese acquired from retail markets in Seoul and Gyeonggi province in Korea from 2005 to 2010 were monitored using a one-step multiplex polymerase chain reaction (PCR) method that involves the amplification of specific soya or maize endogenous genes and the amplification of 35S promoter (p35S) and nopaline synthase terminator (tNOS) for GMO detection. Among the 543 samples, 477 samples were amplified for maize and/or soybean endogenous genes. Although one sausage sample collected in 2008 showed amplification of tNOS, the result was assumed to be false positive based on the results from further tests of other sausage samples of the same brand. Our results demonstrate the absence of GM soya and/or maze of livestock products in the Korean market during 2005-2010. In addition, the one-step multiplex PCR using previously constructed primer sets appears to be useful as a screening method for the detection of GMOs in processed livestock products. However, more specific methods should be established and employed to detect the event-specific GM gene for positive reaction samples by screening tests in processed livestock products.

• Journal of Genetic Medicine
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• v.17 no.1
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• pp.27-33
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• 2020

Purpose: Duchenne muscular dystrophy (DMD) is the most common lethal muscular dystrophy and is caused by the genetic variants of DMD gene. Because DMD is X-linked recessive and shows familial aggregates, prenatal diagnosis is an important role in the management of DMD family. We present our experience of prenatal molecular diagnosis and carrier detection based on multiplex polymerase chain reaction (PCR), multiplex ligation-dependent probe amplification (MLPA), and linkage analysis. Materials and Methods: During study period, 34 cases of prenatal diagnosis and 21 cases of carrier detection were performed at the Seoul National University Hospital. Multiplex PCR and MLPA was used to detect the exon deletions or duplications. When the DMD pathogenic variant in the affected males is unknown and no DMD pathogenic variant is detected in atrisk females, linkage analysis was used. Results: The prenatal molecular diagnosis was offered to 34 fetuses. Twenty-five fetuses were male and 6 fetuses (24.0%) were affected. Remaining cases had no pathogenic mutation. We had 24 (80.0%) cases of known proband results; exon deletion mutation in 19 (79.2%) cases and duplication in 5 (20.8%) cases. Linkage analysis was performed in 4 cases in which 2 cases (50.0%) were found to be affected. In the carrier testing, among 21 cases including 15 cases of mother and 6 cases of female relative, 9 (42.9%) cases showed positive results and 12 (57.1%) cases showed negative results. Conclusion: Prenatal molecular diagnosis and carrier detection of DMD are effective and feasible. They are useful in genetic counseling for DMD families.

• Journal of Genetic Medicine
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• v.9 no.1
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• pp.11-16
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• 2012

Purpose: To determine a method to improve the efficacy and accuracy of preimplantation genetic diagnosis (PGD) - polymerase chain reaction (PCR), we compared hot start PCR and conventional multiplex nested PCR. Materials and Methods: This study was performed with single lymphocyte isolated from whole blood samples that were obtained from two couples with osteogenesis imperfecta (OI). We proceeded with conventional multiplex nested PCR and hot start PCR in which essential reaction components were physically removed, and we compared the amplification rate, allele dropout rate and nonspecific products. Afterward, we used selective method for PGD. Results: In the two couples, the respective amplification rate were 93.5% and 80.0% using conventional multiplex nested PCR and 95.5% and 92.0% using hot start PCR. The respective mean allele dropout rates for the two couples were 42.0% and 14.0% with conventional multiplex nested PCR and 36.0% and 6.0% with hot start PCR. Conclusion: The results demonstrate that the hot start PCR procedure provides higher amplification rates and lower allele dropout rate than the conventional method and that it decreased the nonspecific band in multiplex nested PCR. The hot start method is more efficient for analyzing a single blastomere in clinical PGD.

• 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.

• Clinical and Experimental Pediatrics
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• v.53 no.3
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• pp.397-407
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• 2010

Purpose : The F9 gene is known to be the causative gene for hemophilia B, but unfortunately the detection rate for restriction fragment length polymorphism-based linkage analysis is only 55.6%. Direct DNA sequencing can detect 98% of mutations, but this alternative procedure is very costly. Here, we conducted multiplex polymerase chain reactions (PCRs) and conformation sensitive gel electrophoresis (CSGE) to perform a screened DNA sequencing for the F9 gene, and we compared the results with direct sequencing in terms of accuracy, cost, simplicity, and time consumption. Methods : A total of 27 unrelated hemophilia B patients were enrolled. Direct DNA sequencing was performed for 27 patients by a separate institute, and multiplex PCR-CSGE screened sequencing was done in our laboratory. Results of the direct DNA sequencing were used as a reference, to which the results of the multiplex PCR-CSGE screened sequencing were compared. For the patients whose mutation was not detected by the 2 methods, multiplex ligation-dependent probe amplification (MLPA) was conducted. Results : With direct sequencing, the mutations could be identified from 26 patients (96.3%), whereas for multiplex PCRCSGE screened sequencing, the mutations could be detected in 23 (85.2%). One patient's mutation was identified by MLPA. A total of 21 different mutations were found among the 27 patients. Conclusion : Multiplex PCR-CSGE screened DNA sequencing detected 88.9% of mutations and reduced costs by 55.7% compared with direct DNA sequencing. However, it was more labor-intensive and time-consuming.