• Journal of Life Science
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• v.22 no.1
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• pp.110-116
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• 2012

Nuruk plays a significant role in the flavor and quality of Takju and Yakju, which are produced through saccharification and alcohol fermentation by various microorganisms. In this study, we identified microbial strains isolated from a plate count and PCR-denaturing gradient gel electrophoresis (DGGE) analysis targeting the 16S and 28S rRNA genes, in order to characterize bacterial and fungal diversity in Sansung Nuruk. The numbers of bacteria and fungi in Nuruk were $1.5{\times}10^9$ CFU/g and $2.2{\tims}10^8$ CFU/g, respectively. The 16S rRNA gene sequence indicated that the predominant bacteria in the isolates and PCR-DGGE profile of Nuruk were Kocuria spp., Pantoea spp., Lactobacillus spp., Pediococcus spp., Weissella spp., Staphylococcus spp., endophytic bacterium, uncultured Gamma-proteobacteria, uncultured Cyanobacteria, and Actinobacteria. Dominant bacteria from the PCR-DGGE profile were Pediococcous pentosaceus and uncultured Cyanobacteria. The 28S rRNA gene sequence indicated the predominant fungi in the isolates and PCR-DGGE profile to be Trichomonascus spp. Pichia spp., Torulaspora spp., Wickerhamomyces spp., Sacharomycopsis spp., Lichtheimia spp., Mucor spp., Rhizopus spp. Aspergillus spp., and Cladosporium spp. Dominant fungi from the PCR-DGGE profile were Pichia kudriavzevii and Aspergillus oryzae. The PCR-DGGE technique was used for the first time in this study to assess a microbial community in Nuruk and proved to be an effective protocol for profiling microbial diversity.

• Food Engineering Progress
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• v.15 no.4
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• pp.370-375
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• 2011

In this study the polymerase chain reaction (PCR) combined with denaturing gradient gel electrophoresis (DGGE) was evaluated as a method permitting the rapid detection of pathogens in fresh originally grown vegetables. A universal primer (341GCf/534r) was selected for its ability to amplify the V3 region of 16S-rRNA genes in their target pathogens (Salmonella typhimurium, Pseudomonas fluorescens, Bacillus cereus, Listeria monoytogenes, Staphyloocus aureus, E. coli). The 194 bp fragments in PCR were successfully duplicated as expected. The amplified fragments of the same size from six different pathogens also showed good separation upon DGGE. The detection limit of PCR-DGGE for six pathogens in fresh-cut lettuces were over $10^{5}$ CFU/g when sampled by stomaching. However, when the sampling method was changed from stomaching to shaking, the detection limit of six pathogens in organic vegetables was shown to increase by over $10^{1}$ CFU/g, but only those of B. cereus were over $10^{3}$ CFU/g. Therefore, PCR-DGGE was shown to be a reliable method for the detection of pathogens in fresh-cut vegetables.

• Journal of Microbiology
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• v.41 no.2
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• pp.157-160
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• 2003

To estimate genetic relationships within the genus Rhizopus, genetic variations in 20 strains were investigated by DGGE and PCR-RFLP of rDNA ITS region (ITSI, ITS2,5.8S). The size of the amplified products showed the interspecific polymorphisms, 650 bp,700 bp, and 900 bp. The DGGE approach allowed the separation of PCR amplicons of the same length according to their sequence variations. When the rDNA ITS region was digested with six restriction enzymes, 20 strains were classified into five RFLP haplotypes. The range of similarity between the 20 strains by PCR-RFLP was 42.3-100%. Based on the results of DGGE aud PCR-RFLP, the 20 strains were divided into four groups, R. oryzae, R. stolonifer, R. microsporus and R. homothallicus. Further study of R. homothallicus is required.

• Journal of Wetlands Research
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• v.14 no.3
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• pp.419-428
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• 2012

Taxonomic identification of phytoplankton has been a difficult task, even for the experienced taxonomist. Many non-descript, yet abundant, phytoplanktons do exist without distinguishing features which cause difficulties in morphological identification. Using PCR(polymerase chain reaction)-DGGE(denaturing gradient gel electrophoresis)method, which is known to be a powerfulfingerprinting technique to analyze diversity and dynamics of microbial populations, this study aimed to find the way to overcome the limitation of morphological identification. As a result, a total of 46 bands from samples in five lakes were detected in September and 27 bands in November. Fingerprinting results showed convenient and comparative analyses among each sampling site. In this study, PCR-DGGE method was used to figure out diversity and dynamics of plankton community in the lakes of North-Han River system. Also, the possibility of DGGE technique as an identification tool for phytoplankton was estimated.

• Journal of Life Science
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• v.22 no.2
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• pp.232-238
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• 2012

Kumjungsansung-Makgeolli$^{(R)}$ is a traditional Korean rice wine that is fermented from traditional nuruk and rice. In this study, we performed the PCR-denaturing gradient gel electrophoresis (DGGE) analysis targeting the 16S and 28S rRNA genes to characterize bacterial and fungal diversity during Makgeolli fermentation. The predominant bacteria in the PCR-DGGE profile during Makgeolli fermentation were Lactobacillus spp. (Lactobacillus curvatus, L. kisonensis, L. plantarum, L. sakei, and L. gasseri), Pediococcus spp. (P. acidilactici, P. parvulus, P. agglomerans, and P. pentosaceus), Pantoea spp. (P. agglomerans and P. ananatis), and Citrobacter freundii; these were identified on the base of analysis of 16S rRNA gene sequences. The dominant bacterium during Makgeolli fermentation was L. curvatus. The predominant fungi in PCR-DGGE profile during Makgeolli fermentation were Pichia kudriavzevii, Saccharomyces cerevisiae, Asidia idahoensis, Kluyveromyces marxianus, Saccharomycopsis fibuligera, and Torulaspora delbrueckii, and these were identified on the basis of analysis of 28S rRNA gene sequences. The dominant fungal species during Makgeolli fermentation changed from P. kudriavzevii at 0-2 days incubation to S. cerevisiae at 3-6 days incubation. This study suggests that PCR-DGGE analysis could be a suitable tool for the understanding of microbial diversity and structure during Makgeolli fermentation.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.12
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• pp.1887-1894
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• 2007

In this paper, a rapid and reliable gene-targeted species-specific polymerase chain reaction (PCR) technique based on a two-step process was established to identify bifidobacteria in dairy products. The first step was the PCR assay for genus Bifidobacterium with genus specific primers followed by the second step, which identified the species level with species-specific primer mixtures. Ten specific primer pairs, designed from nucleotide sequences of the 16-23S rRNA region, were developed for the Bifidobacterium species including B. angulatum, B. animalis, B. bifidum, B. breve, B. catenulatum, B. infantis, B. longum, B. minimum, B. subtile, and B. thermophilum. This technique was applied to the identification of Bifidobacterium species isolated from 6 probiotic products, and four different Bifidobacterium spp. (B. bifidum, B. longum, B. infantis, and B. breve) were identified. The findings indicated that the 16S-23S rDNA gene-targeted species-specific PCR technique is a simple and reliable method for identification of bifidobacteria in probiotic products. PCR combined with Denaturing Gradient Gel Electrophoresis (DGGE) for identification of the bifidobacteria was also evaluated and compared with the gene-targeted species-specific technique. Results indicated that for fermented milk products consistency was found for both species-specific PCR and PCR-DGGE in detecting species. However, in some lyophilized products, the bands corresponding to these species were not visualized in the DGGE profile but the specific PCR gave a positive result.

• Microbiology and Biotechnology Letters
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• v.41 no.3
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• pp.292-299
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• 2013

In this study, we analyzed the changes in fungal populations of red-pepper fields employing eco-friendly farming methods, such as microbial agents and crop rotation, by using polymerase chain reactions coupled with denaturing gradient gel electrophoresis (PCR-DGGE). Primer specific for fungi were used to determine the contribution of domains to the microbial community. Analysis of planted and non-planted soil samples applying PCR-DGGE technology offered evaluation of long-term patterns in fungal species richness. To evaluate the stability of DGGE patterns from different soils, comparison of planted and non-planted soil samples were compared using PCR-DGGE. The number of DNA fragments obtained from all planted soil samples by DGGE separation was far greater (14 to 15 bands) than that of the non-planted soil samples (3 to 4 bands). In addition, 14 bands were observed from crop continuation soil treated with agrochemicals and 18 bands from crop rotation soil treated with microbial agents. The PCR-DGGE analysis suggests that the use of crop rotation and microbial agents benefits the fungal community more than crop continuation using agrochemicals. These results indicate that crop rotation with microbial agents was better able to support beneficial organisms, enable more effective biological control and maintain a healthier balance of nutrients, organic matter and microorganisms.

• Korean Journal of Environment and Ecology
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• v.24 no.3
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• pp.318-324
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• 2010

The aim of this study is to examine feeding habits of the Korean water deer(Hydropotes inermis argyropus) from its rumen contents using a PCR-DGGE method. For this study, rumen contents were collected from water deer causalities by natural death or road-kill in two different sites(Cheorwon, Gangwon province and the Eastern part of Jeonnam province). DNA was extracted from rumen contents of a total of 44 individuals. Two primers, rbcLZ1aF(GC) and rbcL19bR, were used for PCR amplifications of ribulose-1,5-bisphosphate carboxylase large subunit (rbcL) gene. Among 44 samples, twenty-nine samples were successfully amplified by PCRs. The 29 PCR products of partial rbcL gene were applied for PCR-DGGE. Totally, six families of plants were detected from the diet analyses. Five families of plants were found in Cheorwon, Gangwon province, but only three families of plants were found in the Eastern part of Jeonnam province. The PCR-DGGE method will provide us with a potential tool to study feeding habits of ungulates including water deer, even though our results failed to identify the prey plants at the level of species.

• Journal of Microbiology and Biotechnology
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• v.20 no.11
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• pp.1592-1596
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• 2010

To verify the dominance of microorganisms in wastewater biological treatment, PCR-DGGE (denaturing gradient gel electrophoresis) was performed as a supplementary support method for screening of the dominant microorganisms from activated sludge. Results suggest that the dominant microorganisms in activated sludge are primarily responsible for strengthening its effectiveness as a biological treatment system, followed by the non-main dominant microorganisms, whereas the non-dominant microorganisms showed no effects. The degree of microbial abundance present on the profile of PCR-DGGE was in line with the treatment efficiency of augmented activated sludge with isolated cultures, suggesting that PCR-DGGE can be used as an effective supplementary method for verifying culturable dominant microorganisms in activated sludge of coking wastewater.

• Journal of Korean Society of Environmental Engineers
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• v.28 no.7
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• pp.752-756
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• 2006

The independent anoxic reactor was introduced in biological aerated filters as the regulation of water quality requirement, especially total nitrogen, had been strengthened. The process studied in this work was upflow $Biobead^{(R)}$ process which was used commercial invented for removal of organic materials and nitrification. For the purpose of evaluating the independent anoxic reactor, PCR-DGGE, of the molecular biological methods, was performed. Two types of nitrite reductase genes were selected. One is nirS represented cytocrome $cd_1$ nitrite reductase gene and the other is nirK represented Cu-containing nitrite reductase gene. Denitrifier community in the independent anoxic reactor was analyzed with PCR-DGGE using these two denitrifying functional genes. As the result of the PCR, only nirS gene was detected between nirS and nirK. With the result of the DGGE, specific bands became strong, as the operating days were longer, nitrate loading rate was increased. otherwise those of the initial activated sludge showed various bands. In the consequence of the sequence of DGGE bands, various denitrifiers were sequenced in the initial activated sludge, while specific denitrifiers like alcaligenes faecalis were predominant in the anoxic reactor. Consequently, introduction of the independent anoxic reactor made it possible to achieve 96% denitrification efficiency, and was proper for the modification of BAF process.