• International Journal of Oral Biology
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• 제46권3호
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• pp.140-145
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• 2021

This study aimed to develop Lautropia mirabilis-specific quantitative real-time polymerase chain reaction (qPCR) primers based on the sequence of DNA-directed RNA polymerase subunit beta gene. The PrimerSelect program was used in designing of the qPCR primers, RTLam-F4 and RTLam-R3. The specificity of the qPCR primers were performed by conventional PCR with 37 strains of 37 oral bacterial species, including L. mirabilis. The sensitivity of the primers was determined by qPCR with the serial dilution of purified genomic DNA of L. mirabilis KCOM 3484, ranged from 4 ng to 4 fg. The data showed that the qPCR primers could detect only L. mirabilis strains and as little as 40 fg of genome DNA of L. mirabilis KCOM 3484. These results indicate that this qPCR primer pair (RTLam-F4/RTLam-R3) may be useful for species-specific detection of L. mirabilis in epidemiological studies of oral bacterial infectious diseases such as periodontal disease.

• 한국미생물·생명공학회지
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• 제22권6호
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• pp.577-583
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• 1994

To classify Lactobacillus casei strains on the basis of difference in their chromosomal DNA sequence, we have performed polymerase chain reactions on their chromosomal DNA by using random primers, and followed by analyzing randomly amplified polymorphic DNA fragments. We also developed a mini-preparative method to isolate PCR-grade chromosomal DNA from Lacto- bacillus casei strains within 3 hours. Based on RAPD pattems by polymerase chain reactions with degenerated random primers, 4 Lactobacillus casei strains and 2 mutant strains were successfully discriminated. Results were very sensitive, strain-specific and reproducible. It was also reliable. These results suggest that RAPD may be applied efficiently for the identification of several Lactoba- cillus casei strains.

• International Journal of Oral Biology
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• 제46권4호
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• pp.155-159
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• 2021

This study aimed to develop strain-specific polymerase chain reaction (PCR) primers to detect Fusobacterium hwasookii KCOM 1249^T, F. hwasookii KCOM 1253, F. hwasookii KCOM 1256, F. hwasookii KCOM 1258, and F. hwasookii KCOM 1268 on the basis of nucleotide sequences of a gene specific to each strain. The unique genes for each F. hwasookii strain were determined on the basis of their genome sequences using Roary. The strain-specific PCR primers based on each strain-specific gene were designed using PrimerSelect. The specificity of each PCR primer was determined using the genomic DNA of the 5 F. hwasookii strains and 25 strains of oral bacterial species. The detection limit and sensitivity of each strain-specific PCR primer pair were determined using the genomic DNA of each target strain. The results showed that the strain-specific PCR primers correspond to F. hwasookii KCOM 1249^T, F. hwasookii KCOM 1253, F. hwasookii KCOM 1258, F. hwasookii KCOM 1256/F. nucleatum subsp. polymorphum KCOM 1260, or F. hwasookii KCOM 1268/Fusobacterium sp. oral taxon 203 were developed. The detection limits of these strain-specific PCR primers ranged from 0.2 to 2 ng of genomic DNA for each target strain. The results suggest that these strain-specific PCR primers are valuable in quality control for detecting specific F. hwasookii strains.

• 한국어업기술학회:학술대회논문집
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• 한국어업기술학회 2001년도 춘계 수산관련학회 공동학술대회발표요지집
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• pp.332-333
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• 2001

Genomic DNA was extracted from the blood of the freshwater crucian carp(Carassius carassius) from Kunsan in Korea, representing genetic similarity by polymerase chain reaction amplification of DNA as twelve of arbitrary primers. The electrophoretic analysis of polymerase chain reaction-random amplified polymorphic DNAs(PCR-RADP) products showed the high levels of similarity between different individuals in crucian carp.

• International Journal of Oral Biology
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• 제44권3호
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• pp.96-100
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• 2019

The purpose of this study was to develop Peptoniphilus mikwangii-specific quantitative real-time polymerase chain reaction (qPCR) primers based on the 16S ribosomal RNA (16S rDNA) gene. The specificity of the primers was determined by conventional PCR using 29 strains of 27 oral bacterial species including P. mikwangii. The sensitivity of the primers was determined by qPCR using the purified genomic DNA of P. mikwangii KCOM $1628^T$ (40 ng to 4 fg). The data showed that the qPCR primers (RTB134-F4/RTB134-R4) could detect P. mikwangii strains exclusively and as little as 40 fg of the genomic DNA of P. mikwangii KCOM $1628^T$. These results suggest that the developed qPCR primer pair can be useful for detecting P. mikwangii in epidemiological studies of oral bacterial infectious diseases.

• 한국식물병리학회지
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• 제12권2호
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• pp.191-196
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• 1996

위축, 황화, 총생 증상 등 전형적인 병징을 나타내는 phytoplasma에 감염된 식물에서 phytoplasma만을 특이적으로 검출하기 위하여 polymerase chain reaction(PCR) 방법을 이용하였다. Phytoplasma의 16S rRNA gence의 DNA 단편을 증폭하기 위하여 1.4 kb primers (forward, 5` -GTTGATCCTGGCTCAGGATT-3` 와 reverse, 5` -AACCCCGAGAACGTATTCACC -3`)를 사용하여 증폭한 결과, phytoplasma에 이병된 오동나무, 라일락 및 미역취에서는 약 1.4 kbp의 위치에서 특이 band가 검출되었으나 control로 사용한 건전주에서는 어떠한 band 검출되지 않았다. 위의 결과를 재확인 하기 위하여 약 0.5 kb의 primers(forward, 5` -ACGAAAGCGTGGGGAGCAAA-3` 와 reverse, 5` -GAAGTCGAGTTGCAGACTTC-3`)를 사용하여 증폭한 결과, 0.5 kb의 위치에서 특이 band가 검출되었으나 control로 사용한 건전주에서는 어떠한 band도 검출되지 않았다. Phytoplasma에 이병된 식물의 PCR 반응산물을 제한효소인 AluI으로 처리하 sruf과, 오동나무와 라일락에서는 동일한 band pattern을 나타내어 서로 유연관계가 가까운 phytoplasma인 것으로 생각되며, 미역취에서는 이들과는 다른 band pattern을 나타내어 오동나무와 라일락의 phytoplasma와는 유연관계가 먼 것으로 추측된다.

• 한국동물위생학회지
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• 제23권3호
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• pp.227-233
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• 2000

Salmonella enteritidis is the most prevalent etiologic agents of foodborne acute gastroenteritis. Direct isolation and identification of S enteritidis are time consuming work and not so highly sensitive. This study was conducted to develop for the specific detection of S enteritidis using polymerase chain reaction(PCR). PCR primers were selected to amplify a 351-base pair(bp) DNA fragment from the salmonella plasmid virulence A(spv A) gene of S enteritidis. With the primers, 351 bp DNA products were amplified from S enteritidis but not from other B, D, Cl serogroup Salmonella spp. It was sensitive to detect up to 40 pg of template DNA by agarose gel electrophoresis. This PCR assay is very rapid and specific method and less time consuming than the standard bacteriological methods.

• 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.

• 한국신재생에너지학회:학술대회논문집
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• 한국신재생에너지학회 2005년도 제17회 워크샵 및 추계학술대회
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• pp.631-633
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• 2005

For biohydrogen production, hydrogenase is a key enzyme. In the present work we performed search of [NiFe]-hydrogenases from hydrogen producing microorganisms using degenerate polymerase chain reaction (PCR) strategy. Degenerate primers were designed from the conserved region of [NiFe]-hydrogenase group I especially on structural genes encoding for catalytic subunit of [NiFe]-hydrogenase from bacteria producing hydrogen. Most of [NiFe]-hydrogenase (group I) are expressed via complex mechanism with aid of auxiliary protein and localized through twin-arginine translocation pathway. [NiFe]-hydrogenase is composed of large and small subunits for catalytic activity. It is known that only small subunit has signal peptide for periplasmic localization and large & small subunitscome together before localization. During this process, large subunit is treated by endopeptidase for maturation. Based on these information we used signal peptide sequence and C-terminal of large subunit by recognized by endopeptidase as templates for degenerate primers. About 2,900 bp of PCR products were successfully amplified using the designed degenerate primers from genomic DNAs of several microorganisms. The amplified PCR products were inserted into T-vector and then sequenced to confirm.

• 한국식물병리학회지
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• 제14권4호
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• pp.308-313
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• 1998

Reverse transcription and polymerase chain reaction (RT-PCR) techiniques were used to identification and differentiation of cucumber mosaic virus isolated from Forsythia koreana (CMV-Fk). RT-PCT used by two set of 20-mer primers one was CMV-common primers and another was CMV subgroup I-specific primers designed in a conserved region of the 3' end of CMV RNA3, amplified about 490 bp and 200 bp DNA fragments from CMV-Fk, respectively. CMV could be detected by RT-PCR at a dilution as low as 10-4 in forsythia crude sap extracts. Restriction enzyme analysis of RT-PCR products using EcoRI and MspI showed that CMV-Fk belonged to CMV subgroup I. But, analysis of RNA fingerprinting by arbitrarily primed polymerase chain reaction (RAP-PCR) showed heterogeneity of RNA3 between CMV-Fk and CMV-Y as a member of subgroup I.