• Applied Biological Chemistry
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• v.46 no.4
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• pp.344-347
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• 2003

A method using PCR was developed for the monitoring of genetically modified soybean (GMS) and GMS derived foods utilized in the market. We designed 3 pairs of specific oligonucleotide primers based on epsps and pat inserted in GMS and ferritin gene as internal standards. Template DNAs isolated from soybean and processed foods were used for multiplex PCR with 3 primer sets. PCR, used with specific primer sets for GMS detection, showed the amplified DNA fragments with GMS template DNA. In this study, GMS containing epsps was detected from soy processed foods manufactured before GM food labeling system, however, GMS containing epsps or pat was not detected from soy processed foods manufactured after GM food labeling system.

• Journal of Applied Biological Chemistry
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• v.58 no.4
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• pp.383-387
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• 2015

A duplex polymerase chain reaction (PCR) detection method was developed to authenticate the use of cat and rabbit in food and to prevent unlawful distribution of illegally butchered meat in both domestic and imported food market. Species-specific primers were designed targeting mitochondrial cytochrome b gene. The sizes of PCR products were 191 bp for cat and 101 bp for rabbit, which were relatively small for better application of the detection method on processed foods. Specificities of primers were verified using 21 animal species including cat and rabbit. Limit of detection was examined by serial dilution of the sample DNA and confirmed as 0.005 ng for rabbit and 0.0005 ng for cat using Bioanalyzer. The developed duplex PCR method showed specificity and sensitivity in the identification of two target species.

• Journal of Microbiology and Biotechnology
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• v.17 no.3
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• pp.511-515
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• 2007

White-spot syndrome virus (WSSV) is a devastating, infectious virus affecting shrimp. Although sensitive techniques involving PCR have been developed to assist farmers in screening shrimp (brood stock) for WSSV prior to stocking ponds, such practices have not yet been applied in Korea. Despite the rationality of implementing screening, there has been some doubt as to whether the stocking of WSSV-PCR-negative fly epidemiologically decreases white-spot disease outbreaks. Here, we report a retrospective analysis of data from shrimp farms in the western coast of Korea where WSSV-PCR-negative brood stocks were used to stock rearing ponds. A total of 366 shrimp from Heuksan Island were sampled for WSSV with PCR. Of the tested shrimp, 7.2% (28 brood stocks) were identified as WSSV positive; only WSSV-PCR-negative shrimp were used for brood stocks. Total unit production (final shrimp production/ the area of the ponds) was higher, at 1.96, in ponds where WSSV-PCR-negative shrimp were used, as compared with 1.02 in other ponds in Korea in 2004. This retrospective analysis of WSSV in Korea may be useful to the shrimp aquaculture industry, suggesting a testable hypothesis that may contribute to the eventual control of WSSV outbreaks.

• Clinical and Experimental Reproductive Medicine
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• v.32 no.4
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• pp.293-300
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• 2005

연구목적: 단일 유전자 이상에 대한 착상전 유전진단을 성공적으로 시행하기 위해서는 효과적이고 신뢰도가 높은 PCR 방법의 확립이 중요하다. 본 연구에서는 alkaline lysis와 duplex nested PCR 방법을 단일 림프구와 할구의 유전자 분석에 적용하여 그 효용성을 확인하고자 하였다. 재료 및 방법: 단일 유전자의 이상이 확인된 Duchenne muscular dystrophy (DMD), ornithine transcarbamylase (OTC) 결핍증과 epidermolysis bullosa (EB) 가계의 대상자들에서 채취한 단일 림프구와 공여 받은 배아의 할구를 이용하여 각각 PCR, restriction fragment length polymorphism (RFLP)와 direct DNA sequencing 분석을 시행하였다. 이러한 분석에서 유전자 증폭률 (amplification rate)과 두개의 allele 중에서 하나의 allele이 증폭되지 않는 allele drop-out (ADO) 빈도에 대해 살펴보았다. 결 과: 단일 림프구와 할구를 이용한 PCR 방법의 유전자 증폭률은 DMD에서 91.1%와 81.8%, OTC 결핍증에서 96.0%와 78.1%, EB에서 91.3%와 90.0%를 각각 나타냈으며, ADO 빈도는 OTC 결핍증에서 13.3%, EB에서 16.8%로 관찰되었다. 결 론: 본 연구에서 적용한 alkaline lysis와 duplex nested PCR 방법은 단일 유전자에 대한 착상전 유전진단에 성공적으로 적용할 수 있는 방법으로 생각되며, ADO 빈도를 최소화할 수 있는 효율적인 방법의 개발에 대한 지속적인 연구가 필요하다.

• Microbiology and Biotechnology Letters
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• v.36 no.2
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• pp.96-100
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• 2008

Diversity of lactic acid bacteria involved in 5 Korean fermented vegetables (Cot kimchi, Dongchimi, Baechu kimchi, Oisobagi, and Chonggak kimchi) was investigated using PCR-based method. PCR primer pairs targeted the recA gene were used for the detection of 7 species of lactic acid bacteria mainly found in kimchi and Lactobacillus acidophilus involved in dairy fermentation. Lactobacillus plantarum and Lactobacillus sakei were detected in all samples tested but Lactobacillus paraplantarum, Lactobacillus pentosus, and Lb. acidophilus were not detected. Lactobacillus brevis and Leuconostoc citreum were detected only from Baechu kimchi and Leuconostoc mesenteroides was detected from Got kimchi, Dongchimi, Baechu kimchi, and Oisobagi. The difference of detected species from fermented vegetables may be originated from the difference of main materials. Lb. plantarum and Lb. sakei are supposed to be broadly involved in Korean fermented vegetables.

• Journal of Ginseng Research
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• v.33 no.1
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• pp.55-58
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• 2009

Generally, the direct sequencing through PCR is faster, easier, cheaper, and more practical than clone sequencing. Frequently, standard PCR amplification is usually interpreted by mispriming internal or external regions of the target template. Normally, DNA fragments were eluted from the gel using Gel extraction kit and subjected to direct sequencing or cloning sequencing. Cloning sequencing has often troublesome and needs more time to analyze for many samples. Since touchdown (TD) PCR can generate sufficient and highly specific amplification, it reduces unwanted amplicon generation. Accordingly, TD PCR is a good method for direct sequencing due to amplifying wanted fragment. In plants the 5S-rRNA gene is separated by simple spacers. The 5S-rRNA gene sequence is very well-conserved between plant species while the spacer is species-specific. Therefore, the sequence has been used for phylogenetic studies and species identification. But frequent occurrences of spurious bands caused by complex genomes are encountered in the product spectrum of standard PCR amplification. In conclusion, the TD PCR method can be applied easily to amplify main 5S-rRNA and direct sequencing of panax ginseng cultivars.

• Korean Journal of Veterinary Research
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• v.42 no.4
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• pp.513-522
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• 2002

Antigenic ompC genes of S. gallinarum, S. pullorum and S. dublin were characterized among Salmonella spp. isolated from chickens and other animals to identify genetic variation. Salmonella ompC gene fragment (1,027 bp) was amplified by PCR and the amplicons were cloned for comparison of nucleotide sequences. The identity of the sequences between S. gallinarum and S. pullorum, S. gallinarum and S. dublin, S. pullorum and S. dublin was 99.8%, 97.6% and 97.8%, respectively. Also, we found that ompC has some diversity between S. gallinarum and S. pullorum, and other Salmonella spp. which may be useful to type the organisms. Similar to diagnosis in other organisms, the TaqMan PCR method can be applied to rapid and accurate diagnosis of salmonellosis in chickens and other animals. We designed PCR primers and TaqMan probe for flagellin gene (fliC) for detection of Salmonella spp. by TaqMan PCR. The TaqMan PCR method was 10,000 times more sensitive than conventional PCR.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.7
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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.

• Biomedical Science Letters
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• v.21 no.4
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• pp.227-232
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• 2015

Epstein-Barr virus (EBV) has a pathogenic role in several lymphomas including diffuse large B-cell lymphoma (DLBCL). In this study, we detected EBV from formalin-fixed paraffin embedded (FFPE) tissues of DLBCL patients by RT-PCR and compared the sensitivity of the RT-PCR method to in situ hybridization (ISH) method. The RNA was extracted from 91 FFPE samples with DLBCL and amplified with primers targeting EBV-encoded small RNA (EBER) by RT-PCR. When using the RT-PCR method, 13 of 91 patients (14.3%) were positive and among these 13 cases, 7 cases (7.7%) were from > 50-year-old patients that is classified as EBV positive DLBCL of the elderly. In previous results using ISH method, 3 of 91 patients (3.3%) were positive and 2 case (2.4%) were older than 50-year-old. These results indicate that RT-PCR method used in this study shows a higher sensitivity than ISH method. The ratio of male versus female among the EBV positive samples was 1.2:1 with the ratio of male higher. If RT-PCR method having high sensitivity is used simultaneously as well as the ISH method providing the information of the EBV positive cellular location, it is expected that EBV will be more accurately detected.

• Journal of Korean society of Dental Hygiene
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• v.17 no.1
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• pp.13-25
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• 2017

Objectives: The purpose of this study was to compare colony forming unit (CFU) method and multiplex real-time polymerase chain reaction (MRT-PCR) method for accurate quantitative analysis of bacteria. Methods: We compared the CFU method and the MRT-PCR method, which are still used in Korea, for Prevotella intermedius (P. intermedius), a periodontal disease pathogen selected by MRT-PCR, and Streptococcus mutans (S. mutans), a dental caries causative organism. The subjects of this study were 30 patients who visited the C dental hospital. Results: Total microorganisms in MRT-PCR method were significantly higher in both types of bacteria (p<0.05), since DNA of dead bacteria was also analyzed. This was because the periodontal dise(-) anaerobes, and even dead bacteria contain large amounts of toxic substances called LPS in the extracellular membrane, and fimbriae and pili, which are motility structures, still remain as a strong toxic substance in periodontal tissue. Conclusions: Therefore, in terms of the total amount of bacteria found, the MRT-PCR method will be a useful technique for searching all the bacteria in the oral cavity including live bacteria, as well as sterilization.