• Korean Journal of Microbiology
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• v.39 no.1
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• pp.36-39
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• 2003

The rapid and reliable 16S rDNA multiplex polymerase chain reaction (PCR) assay was established to characterize bacterial etiologies of middle ear effusion. These etiologies included Haemophilus influenzae, Moraxella catarrhalis and Streptococcus pneumonia, which were detected in middle-ear effusion (MEE) samples taken from patient with otitis media. A total of 39 MEE samples were aspirated from 26 patients. DNA was extracted from MEE samples, and PCR was done with DNA extracts by using the common primers, which is localized at C4 region in the 16S rDNA gene of all bacterial species, and species-specific primers: (i) Haemophilus-specific primer, (ii) Moraxella- specific primer, and (iii) Streptococcus-specific primer. Among 39 samples tested, 24 (61.5%) were positive for H. influenzae, 10 (25.6%) were positive for M. catarrhalis, 3(7.7%) were positive for S. pneumonia, and 11 (28%) were negative for 165 rDNA multiplex PCR reaction. Nine samples (28.6%) exhibited a mixed infection and were positive for both H. infuenzae and M. catarrhalis. We suggested that 16S rDNA multiplex PCR is a useful method to identify rapidly for rapid identification of the pathogenic bacteria and characterization of bacterial etiologies of middle ear effusion.

• Korean Journal of Veterinary Research
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• v.45 no.2
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• pp.199-205
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• 2005

DNA fingerprint patterns of 8 Brucella reference strains and 15 B. abortus field isolates were characterized by repetitive element sequence-based PCR (rep-PCR) using BOX- and ERIC-primers in this study. AMOS PCR differentiated all Brucella field isolates from B. abortus RB51, a vaccine strain by producing a B. abortus-specific 498 bp band. Rep-PCR using BOX-primer produced 13 to 18 bands with sizes of between 230 and 3,300 bp, and discriminated Brucella strains to the species level except B. canis and B. suis. PCR products amplified with ERIC primers were, however, not appropriate for differentiating the Brucella isolates. DNA fingerprint patterns for all B. abortus field isolates were identical among them and were put on one cluster with B. abortus biovar 1 reference strain in the dendrogram, indicating they were highly clonal. These results suggested that rep-PCR using BOX primer might to be a useful tool for calculating genetic relatedness among the Brucella species and for the study of brucellosis epidemiology.

• Fisheries and Aquatic Sciences
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• v.26 no.11
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• pp.678-688
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• 2023

Flatfish are one of the largest families in the order Pleuronectiformes and are economically important edible marine fish species. However, they have similar morphological characteristics leading to challenges in classifying correctly, which may result in mislabeling and illegal sales, such as fraudulent labeling of processed food. Therefore, accurate identification is important to ensure the quality and safety of domestic markets in Korea. Species-specific primers were prepared from the mainly consumed eleven species of the order Pleuronectiformes. To rapidly identify the 11 flatfish species, a highly efficient, rapid, multiplex polymerase chain reaction (PCR) with species-specific primers was developed. Species-specific primer sets were designed for the mitochondrial DNA cytochrome c oxidase subunit I gene. Species-specific multiplex PCR (MSS-PCR) either specifically amplified a PCR product of a unique size or failed. This MSS-PCR analysis is easy to perform and yields reliable results in less time than the previous Sanger sequencing methods. This technique could be a powerful tool for the identification of the 11 species b the family Pleuronectidae and can contribute to the prevention of falsified labeling and protection of consumer rights.

• Korean Journal of Veterinary Service
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• v.35 no.4
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• pp.289-294
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• 2012

Mycoplasmas are highly fastidious bacteria, difficult to culture and slow growing. Infections with Mycoplasma species can cause a variety of problems in living organisms and in vitro cell cultures. In this study, we investigated the usefulness of a genus-specific consensus PCR analysis method to detect Mycoplasma species. The developed consensus primer pairs MycoF and MycoR were designed specifically to amplify the 16S ribosomal RNA gene (rRNA) of Mycoplasma species by the optimized PCR system. The developed consensus PCR system effectively amplified 215 bp of Mycoplasma genus-specific region of 16S rRNA. In conclusion, we recommend this consensus PCR for monitoring Mycoplasma species in animals, human and cell culture system.

• Mycobiology
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• v.31 no.1
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• pp.23-31
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• 2003

With universal primer ITS1-F, the specific DHJ2 primer was developed to detect the Ectomycorrhizal(ECM) root tips in soil and to identify the species of ECM fungi, as based on DNA sequences of rDNA stored in GeneBank of NCBI. This primer was designed with the common sites of rDNA of Amanita and Boletus, and was also designed with several DNA programs provided by NCBI. The DNA fragments synthesized by PCR were calculated to be 1,000 to 1,200 bps of DNA located to 18s to 28s rDNA to contain two variable sites of ITS, indicating much diversities for specific species or ecotypes of ECM fungi. The primer DHJ2 reacted with the genomic DNA's extracted from the tissues of basidiocarp at the rate of 73 of 80 fungi collected produced single bands with a 1,100 bps length. The DNA fragment synthesized with the genomic DNA that extracted from eight ECM tips of Pinus densiflora was confirmed and analysized to the rDNAs of ECM in full sequences, and informed to be a ECM fungal species in the forest.

• Food Science of Animal Resources
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• v.27 no.2
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• pp.209-215
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• 2007

In order to develop a sensitive and reliable method for the species-specific molecular markers, PCR-RFLP assay of the mitochondrial DNA(mt DNA) 12S rRNA gene was exploited for the identification of the origin of animal meat species including cattle, pig, sheep, goat, horse, deer, chicken, duck and turkey. A specific primer pairs were designed, based on the nucleotide sequences of mt 12S rRNA gene, for the amplification of the highly conserved region in the gene of the animal species using PCR-RFLP technique. mt DNA was isolated from meat samples followed by DNA amplification using PCR with the specific primers. PCR amplification produced an approximately 455 bp fragment in each of these animal meats. To distinguish pleat species, the PCR amplicons were digested with restriction endonucleases Tsp5091 and MboI, respectively, which generates distinct RFLP profiles. The DNA profiles digested with Tsp5091 allowed the clear discrimination in the mammalian meat species and the DNA profiles digested with MboI in poultry meat species. Therefore, the PCR-RFLP profiles of mt 12S rRNA gene could be very useful to identify the origin of the raw materials in the raw meats as well as the processed meat products.

• Applied Biological Chemistry
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• v.49 no.3
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• pp.192-195
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• 2006

For the development of a qualitative PCR detection method for genetically modified perilla (Perilla frutescens), perilla species-specific gene, KAS-I (Beta-ketoacyl-ACP synthase I), was selected and validated as suitable for the use as an endogenous reference gene in perilla. Primer specificity was first tested by the means of qualitative PCR analysis. The primer pair Pfru3-F/R amplifying the perilla endogenous gene, KAS-I, gave rise to an amplicon 95 bp. No amplified product was observed when DNA samples from 15 different plants were used as templates. Qualitative PCR detection method was assayed with vitamin E-enriched GM Perilla developed in Korea. For the qualitative PCR detection method, the construct-specific detection primer pairs were constructed. The primer pair TMTO-F/R amplifying the junction region of TMT (${\gamma}$-tocopherol methyltransferase) gene and OCS (Octopine synthase) terminator introduced in GM perilla gave rise to an amplicon 148 bp.

• Journal of Industrial Technology
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• v.28 no.A
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• pp.141-147
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• 2008

Several contaminated bacteria such as Lactobacillus brevis and Pediococcus damnosus in beer production cause beer spoilage by producing off flavours and turbidity. Detection of these organisms is complicated by the strict anaerobic conditions and lengthy incubation times required for their cultivation, consequently there is a need for more rapid detection methods. Recently, two contaminated strains were isolated from vessel of beer production and identified as Lactobacillus species by API kit identificaton as well as 16S-23S ITS sequencing analyses. Two isolated strains were named as Lactobacillus sp. HLA1 and Lactobacillus HLB2, respectively. A polymerase chain reaction (PCR) method was developed for the rapid and specific detection of Lactobacillus sp.. Two sets of primer pairs (HLA1-F/HLA1-R and HLB2-F/HLB2-R) were designed for the amplification of a 1576 base pair (bp) fragment of the HLA1 16S-23S rRNA gene and 1888 bp fragement of the HLB2 16S-23S rRNA. Amplified PCR products were highly specific to detect corresponding bacteria when other contaminated strains were used as PCR templates. However, detection of both strains were limited when $100{\mu}{\ell}$ of cultured samples were mixed with $100m{\ell}$ of beer sample in arbitrary manner. The sensitivity of the assay still needs to be improved for direct detection of the small amounts of bacteria present in beer.

• Journal of Microbiology and Biotechnology
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• v.17 no.5
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• pp.806-811
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• 2007

Bacillus anthracis is a soil pathogen capable of causing anthrax that is closely related to several environmental species, including B. cereus, B. mycoides, and B. thuringiensis. DNA homology studies showed that B. anthracis, B. cereus, B. mycoides, and B. thuringiensis are closely related, with a high sequence homology. To establish a method to specifically detect B. anthracis in situations such as environmental contamination, we initially performed RAPD-PCR with a 10-mer random primer and confirmed the presence of specific PCR bands only in B. anthracis species. One region specific for B. anthracis was cloned and sequenced, and an internal primer set was designed to amplify a 241-bp DNA fragment within the sequenced region. The PCR system involving these specific primer sets has practical applications. Using lyses methods to prepare the samples for PCR, it was possible to quickly amplify the 241-bp DNA segment from samples containing only a few bacteria. Thus, the PCR detection method developed in this study is expected to facilitate the monitoring of environmental B. anthracis contamination.

• Korean Journal of Veterinary Service
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• v.38 no.2
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• pp.101-106
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• 2015

Murine mycoplasmosis, caused by Mycoplasma (M.) pulmonis, is a prominent disease in rodent animals. The aim of this study was to develop a sensitive and specific PCR assay to detect M. pulmonis in animals and to assess the suitability of this assay for the detection of mycoplasmal infection in rats experimentally infected with M. pulmonis. A new PCR assay using the M. pulmonis-specific primer pairs MPul-F and MPul-R was developed. The primers and probe for the assay were designed from regions in the 16S rRNA gene that are unique to M. pulmonis. The novel PCR assay was very specific and sensitive for M. pulmonis, detecting the equivalent of 5 pg of target template DNA. It detected only M. pulmonis and no other Mycoplasma species or other bacterial species. The newly developed PCR assay also effectively detected M. pulmonis infection in rats. These results suggest that this PCR assay using M. pulmonis-specific primer pairs of MPul-F and MPul-R will be useful and effective for monitoring M. pulmonis infection in animals.