• 한국잠사학회:학술대회논문집
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• 한국잠사학회 2003년도 International Symposium of Silkworm/Insect Biotechnology and Annual Meeting of Korea Society of Sericultural Science
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• pp.138-140
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• 2003

We report the development of a multiplex PCR-based procedure for rapid, sensitive, and specific detection of entomopathgenic microspordia and differentiation of the genus Nosema and Vairimorpha and identification of the vairimorpha necatrix. (omitted)

• 한국수의병리학회:학술대회논문집
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• 한국수의병리학회 2003년도 추계학술대회초록집
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• pp.27-27
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• 2003

Porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV) infections are considered difficult to distinguish clinically and histopathologically. Prompt differentiation between PEDV- and TGEV-associated enteritis would greatly facilitate the management of disease in countries where PEDV and TGEV are epizootic. Rapid differential diagnosis and treatment are crucial to reducing mortality and morbidity from PEDV- and TGEV-induced enteritis in piglets. The objective for this study was to develop a protocol to differentiate between PEDV and TGEV directly from formalin-fixed, paraffin-embedded tissue, using a multiplex reverse transcription-nested polymerase chain reaction (RT-nPCR) assay. (omitted)

• Journal of Genetic Medicine
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• 제9권1호
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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.

• 한국산학기술학회논문지
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• 제14권3호
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• pp.1191-1196
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• 2013

임상검체에서 분리되는 그람음성 막대균의 제 3세대 cephalosporin에 대한 내성율의 증가는 임상적으로 심각한 문제가 되고 있다. 3세대 cephalosporin 및 monobactam계 항균제에 대한 내성은 주로 Extended-Spectrum ${\beta}$-Lactamase(ESBL)의 생성에 기인한다. 따라서 ESBL 유전자의 정확한 검출은 병원내의 감염경로 파악을 위한 감시 및 역학조사를 위해 필수적이다. 본 연구는 2012년 2월부터 8월까지 대전, 충남, 충북지역의 대학병원으로부터 ESBL 생성 Klebsiella penumoniae 46균주를 분리하여 Clinical and Laboratory Standards Institute (CLSI)에 따라 ceftazidime (CAZ)과 CAZ/clavulanate (CLA)를 이용한 combination disk test (CDT) 방법에 의해 표현형을 조사하고, 유전형 특이 프라이머를 이용한 multiplex PCR을 수행하여 유전형을 검출하였다. CDT 결과 42균주가 ESBL생성균주로 확인되었다. PCR 결과, 46균주 모두 TEM형이었으며, 37균주는 SHV형, 14균주는 CTX-M형으로 나타났으며 10균주가 TEM, SHV, CTX-M 유전자를 모두 가지고 있었다. Multiplex PCR에 의한 유전형 검출 방법은 임상에서 분리한 ESBL생성 K. penumoniae균주의 감별과 검출에 유용한 방법으로 사료된다.

• Asian-Australasian Journal of Animal Sciences
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• 제26권7호
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• pp.916-920
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• 2013

The triploid Pacific oyster, which is produced by mating tetraploid and diploid oysters, is favored by the aquaculture industry because of its better flavor and firmer texture, particularly during the summer. However, tetraploid oyster production is not feasible in all oysters; the development of tetraploid oysters is ongoing in some oyster species. Thus, a method for ploidy verification is necessary for this endeavor, in addition to ploidy verification in aquaculture farms and in the natural environment. In this study, a method for ploidy verification of triploid and diploid oysters was developed using multiplex polymerase chain reaction (PCR) panels containing primers for molecular microsatellite markers. Two microsatellite multiplex PCR panels consisting of three markers each were developed using previously developed microsatellite markers that were optimized for performance. Both panels were able to verify the ploidy levels of 30 triploid oysters with 100% accuracy, illustrating the utility of microsatellite markers as a tool for verifying the ploidy of individual oysters.

• 대한수의학회지
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• 제52권2호
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• pp.75-82
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• 2012

We have developed and optimized a multiplex polymerase chain reaction (mPCR) for simultaneous detection of Brucella, Salmonella and Leptospira with high sensitivity and specificity. Three pairs of oligonucleotide primers were designed to specifically amplify the targeted genes of Salmonella, Leptospira and Brucella species with sizes of 521, 408 and 223 bp, respectively. The mPCR did not produce any nonspecific amplification products when tested against 15 related species of bacteria. The sensitivity of the mPCR was 100 fg for Brucella and 1 pg for both Salmonella and Leptospira species. In the field application, kidney, liver and spleen were collected from wild rats and stray cats and examined by mPCR. The high specificity and sensitivity of this mPCR assay provide a valuable tool for diagnosis and for the simultaneous and rapid detection of three zoonotic bacteria that cause disease in both humans and animals. Therefore, this assay could be a useful alternative to the conventional method of culture and single PCR for the detection of each pathogen.

• Asian Pacific Journal of Cancer Prevention
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• 제16권7호
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• pp.3003-3007
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• 2015

Objective: To explore the application of multiplex nested methylated specific polymerase chain reaction (PCR) in the early diagnosis of epithelial ovarian carcinoma (EOC). Materials and Methods: Serum and fresh tissue samples were collected from 114 EOC patients. RUNX3, TFPI2 and OPCML served as target genes. Methylation levels of tissues were assessed by multiplex nested methylated specific PCR, the results being compared with those for carcinoma antigen 125 (CA125). Results: The serum free deoxyribose nucleic acid (DNA) methylation spectrum of EOC patients was completely contained in the DNA spectrum of cancer tissues, providing an accurate reflection of tumor DNA methylation conditions. Serum levels of CA125 and free DNA methylation in the EOC group were evidently higher than those in benign lesion and control groups (p<0.05). Patients with early EOC had markedly lower serum CA125 than those with advanced EOC (p<0.05), but there was no significant difference in free DNA methylation (p>0.05). The sensitivity, specificity and positive predicative value (PPV) of multiplex nested methylated specific PCR were significantly higher for detection of all patients and those with early EOC than those for CA125 (p<0.05). In the detection of patients with advanced EOC, the PPV of CA125 detection was obviously lower than that of multiplex nested methylated specific PCR (p>0.05), but there was no significant difference in sensitivity (p>0.05). Conclusions: Serum free DNA methylation can be used as a biological marker for EOC and multiplex nested methylated specific PCR should be considered for early diagnosis since it can accurately determine tumor methylation conditions.

• Journal of Plant Biotechnology
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• 제47권4호
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• pp.309-315
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• 2020

Advances in biotechnology have led to progress in crop genetic engineering to improve agricultural productivity. The use of genetically modified (GM) crops has increased, as have consumers' and regulators' concerns about the safety of GM crops to human health, and ecological biodiversity. As such, the identification of GM crops is a critical issue for developers and distributors, and their labeling is mandatory. Multiplex polymerase chain reaction (PCR) has been developed and its use validated for the detection and identification of GM crops in quarantine. Herein, we established a simultaneous detection method to identify four GM maize events. Event-specific primers were designed between the junction region of transgene and genome of four GM maize lines, namely 5307, DAS-40278-9, MON87460, and MON87427. To verify the efficiency and accuracy of the multiplex PCR we used specificity analysis, limit of detection evaluation, and mixed certified reference materials identification. The multiplex PCR method was applied to analyze 29 living, modified maize volunteers collected in South Korea in 2018 and 2019. We performed multiplex PCR analysis to identify events and confirmed the result by simplex PCR using each event-specific primer. As a result, rather than detecting each event individually, the simultaneous detection PCR method enabled the rapid analysis of 29 GM maize volunteers. Thus, the novel multiplex PCR method is applicable for living modified organism volunteer identification.

• 한국임상수의학회지
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• 제23권3호
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• pp.300-307
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• 2006

In the present study, methods of the reverse transcription-polymerase chain reaction(RT-PCR) were evaluated for the rapid detection and differentiation of transmissible gastroenteritis virus(TGEV), porcine epidemic diarrhea virus(PEDV) and rotavirus in piglets suffering from diarrhea. For the purposes, the PCR conditions were first confirmed for the amplification of VP7 gene of rotavirus and N gene of TGEV and PEDV using each specific primers and their annealing temperature. Multiplex RT-PCR methods were further determined to distinguish these viral infections and the results are as follows. For the specific amplification of these viral genes, the reliable PCR condition was determined as 30 cycles of reaction consisting each 1 min of denature at $94^{\circ}C$, annealing at $42^{\circ}C$ and polymerization at $72^{\circ}C$ with 1.0 mM $MgCl_2$. It was able to differentiate these viral infections in the intestines and feces of piglets suffering from diarrhea by duplex PCR for TGEV and PEDV and single PCR for rotavirus with a primer-annealing temperature of $42^{\circ}C$. When the multiplex RT-PCR were undertaken for the field samples, 17 cases of PEDV and 5 cases of rotavirus infections were differential diagnosed in a total of 92 samples of intestines and feces of the piglets with diarrhea.

• 대한임상검사과학회지
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• 제49권1호
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• pp.22-27
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• 2017

The length of hospital stay (LOS) for patients with respiratory virus infections has been reported to depend the virus type and infection severity. However, the impact of co-infections remains unclear. Patients with suspected respiratory virus infections, who visited Dankook University Hospital between December 2006 and February 2014, were included to examine the relationship between co-infections and LOS. Multiplex reverse transcriptase-polymerase chain reactions were used to identify the causative viruses. LOS was analyzed with respect to sex, age, virus, and co-infection. During this period, 5,310 out of the 8,860 patients (59.9%; median age, 1.5 years) were respiratory virus-positive. In respiratory virus-positive patients with single, double, and three-or-more infections, the average LOS was 7.3, 6.7, and 6.6 days, respectively. Longer LOS was observed for older patients and those with human coronavirus OC43 infections compared with adenovirus or respiratory syncytial virus A infections. LOS differed significantly according to age, virus type, and co-infection, but not between double and three or more infections.