• Animal Bioscience
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• 제36권2_spc호
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• pp.364-373
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• 2023

The gastrointestinal (GI) tract of ruminants contains diverse microbes that ferment various feeds ingested by animals to produce various fermentation products, such as volatile fatty acids. Fermentation products can affect animal performance, health, and well-being. Within the GI microbes, the ruminal microbes are highly diverse, greatly contribute to fermentation, and are the most important in ruminant nutrition. Although traditional cultivation methods provided knowledge of the metabolism of GI microbes, most of the GI microbes could not be cultured on standard culture media. By contrast, amplicon sequencing of 16S rRNA genes can be used to detect unculturable microbes. Using this approach, ruminant nutritionists and microbiologists have conducted a plethora of nutritional studies, many including dietary interventions, to improve fermentation efficiency and nutrient utilization, which has greatly expanded knowledge of the GI microbiota. This review addresses the GI content sampling method, 16S rRNA gene amplicon sequencing, and bioinformatics analysis and then discusses recent studies on the various factors, such as diet, breed, gender, animal performance, and heat stress, that influence the GI microbiota and thereby ruminant nutrition.

• Animal cells and systems
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• 제6권2호
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• pp.145-151
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• 2002

Phylogenetic signal present in the mitochondrial 16S ribosomal RNA gene (16S rDNA) and the nuclear large subunit ribosomal RNA gene (28S rDNA) was explored to assess their utility in resolving family level relationships of the superfamily Tephritoidea. These two genes were chosen because they appear to evolve at different rates, and might contribute to resolve both shallow and deeper phylogenetic branches within a highly diversified group. For the 16S rDNA data set, the number of aligned sites was 1,258 bp, but 1,204 bp were used for analysis after excluding sites of ambiguous alignment. Among these 1,204 sites, 662 sites were variable and 450 sites were informative for parsimony analysis. For the 28S rDNA data set, the number of aligned sites was 1,102 bp, but 1,000 bp were used for analysis after excluding sites of ambiguous alignment. Among these 1000 sites, 235 sites were variable and 95 sites were informative for parsimony analysis. Our analyses suggest that: (1) while 16S rDNA is useful for resolving more recent phylogenetic divergences, 28S rDNA can be used to define much deeper phylogenetic branches; (2) the combined analysis of the 16S and 28S rDNAs enhances the overall resolution without losing phylogenetic signal from either single gene analysis; and (3) additional genes that evolve at intermediate rates between the 16S and 28S rDNAs are needed to further resolve relationships among the tephritoid families.

• 한국미생물·생명공학회지
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• 제47권3호
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• pp.401-411
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• 2019

All Idiomarina species are isolated from saline environments; microorganisms in such extreme habitats develop metabolic adaptations and can produce compounds such as proteases with an industrial potential. ARDRA and 16S rRNA gene sequencing are established methods for performing phylogenetic analysis and taxonomic identification. However, 16S-23S ITS is more variable than the 16S rRNA gene within a genus, and is therefore, used as a marker to achieve a more precise identification. In this study, ten protease producing Idiomarina strains isolated from the Peruvian salterns were characterized using biochemical and molecular methods to determine their bacterial diversity and industrial potential. In addition, comparison between the length and nucleotide sequences of a 16S-23S ITS region allowed us to assess the inter and intraspecies variability. Based on the 16S rRNA gene, two species of Idiomarina were identified (I. zobellii and I. fontislapidosi). However, biochemical tests revealed that there were differences between the strains of the same species. Moreover, it was found that the ITS contains two tRNA genes, $tRNA^{Ile(GAT)}$ and $tRNA^{Ala(TGC)}$, which are separated by an ISR of a variable size between strains of I. zobellii. In one strain of I. zobellii (PM21), we found nonconserved nucleotides that were previously not reported in the $tRNA^{Ala}$ gene sequences of Idiomarina spp. Thus, based on the biochemical and molecular characteristics, we can conclude that protease producing Idiomarina strains have industrial potential; only two I. zobellii strains (PM48 and PM72) exhibited the same properties. The differences between the other strains could be explained by the presence of subspecies.

• Journal of Microbiology
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• 제44권5호
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• pp.566-571
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• 2006

An archaeal 16S rRNA gene library was constructed from mangrove soil. Phylogenetic analysis revealed archaea in mangrove soil including the Crenarchaeota (80.4%) and Euryarchaeota (19.6%) phyla. The archaeal community in mangrove soil appears to be a mixture of organisms found in a variety of environments with the majority being of marine origin.

• 미생물학회지
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• 제47권3호
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• pp.268-274
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• 2011

남조세균 Anabaena (Cyanobacteria, Nostocales)는 담수 생태계에서 녹조 현상을 유발하거나 일부 종은 간독소(hepatotoxin)를 갖고 있어 수질관리 차원에서 주목 받아 왔다. 본 연구는 Anabaena RNA polymerase beta subunit (rpoB) 유전자 염기서열을 규명하였으며, 분류학적 분자 마커로 사용하기 위하여 이들 염기서열의 특성을 평가하였다. Anabaena rpoB 유전자는 16S rRNA 유전자와 비교하여 염기 유사도가 낮으며 유전자 변이가 큰 것으로 분석되었으며, 통계적으로 유의한 차이를 보였다(Student t-test, p<0.01). Parsimony 분석을 통해 rpoB 유전자가 4.8배의 속도로 빠르게 진화하는 것으로 파악되었다. 또한 rpoB 유전자 phylogeny 분석에서 16S rRNA tree보다 높은 해상도로 Anabaena 균주를 명확하게 구분해 주었다. 본 연구 결과는 Anabaena의 종 식별, 분자계통 분류, 분자적 검출을 위해 rpoB 유전자가 매우 효과적이라는 것을 제시해 준다.

• Journal of Microbiology and Biotechnology
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• 제20권8호
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• pp.1210-1214
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• 2010

Many bacteria are involved in the fermentation of doenjang, and Bacillus species are known to perform significant roles. Although SDS-PAGE has been frequently used to classify and identify bacteria in various samples, the microbial diversity in doenjang has not yet been investigated. This study aims to determine the identity and distribution of dominant Bacillus species in doenjang using SDS-PAGE profiles of whole-cell proteins and 16S rRNA gene sequencing. Reference Bacillus strains yielded differential SDS-PAGE banding patterns that could be considered to be highly specific fingerprints. Grouping of bacterial strains isolated from doenjang samples by whole-cell protein patterns was confirmed by analysis of their 16S rRNA gene sequences. B. subtilis was found to be the most dominant strain in most of the samples, whereas B. licheniformis and B. amyloliquefaciens were less frequently found but were also detected in several samples. The results obtained in this study show that a combined identification method using SDS-PAGE profiles of whole-cell proteins and subsequent 16S rRNA gene sequence analysis could successfully identify Bacillus species isolated from doenjang.

• 한국어류학회지
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• 제18권2호
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• pp.78-86
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• 2006

한국산 쉬리, Coreuleuciscus splendidus의 종내 집단간 유전자 다양성을 알기 위해 6개 주요강(북한강, 남한강, 금강, 오십천, 낙동강, 섬진강)으로부터 채집된 개체를 대상으로 16S rRNA 유전자와 미트콘드리아 cytochrome b 유전자에 근거하여 비교 분석하였다. 미트콘드리아 cytochrome b 유전자의 657 bp 길이의 염기서열 분석결과, 6개 집단간에 차이는 98.2~99.9%로 나타났으며 지리적으로 격리된 집단간에 높은 유전적 다양성을 보였다. 16S rRNA 유전자는 697 bp의 염기서열을 얻었으며, 종내 변이는 큰 차이가 없이 거의 동일하였다. 16S rRNA 유전자의 6개 집단간에는 97.7%에서 99.7%의 높은 유사성을 보였다.

• 한국미생물·생명공학회지
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• 제35권4호
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• pp.347-351
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• 2007

감자 더뎅이병은 제주도 전 지역에서 발생하는 병해로서 더뎅이병 발생 시 경제적인 손실이 막대한 것으로 알려져 있다. 본 연구에서는 제주도 감자 더뎅이병징이 있는 부위에서 Streptomyces spp.를 분리, 배양한 후, 16S rRNA 유전자를 이용하여 계통분석을 실시하였다. 계통분석 결과 제주도 감자 더뎅이병징이 있는 부위에서 분리된 균은 모두 Streptomyces spp.에 속하였으며, 대부분이 기존에 더뎅이병을 일으키는 Streptomyces spp.로 확인되었다. 그러나 일부의 분리균은 기존에 알려진 감자 더뎅이병원균과는 다르다. 따라서 이들 병원균에 대해서는 지방산과 단백질 분석, 그리고 DNA-DNA 혼성화 등과 같은 보다 많은 연구의 수행을 통하여 새로운 더뎅이병을 일으키는 Streptomyces spp.인지, 또는 아직까지 명명되지 않은 Streptomyces spp.인지를 확인해야 할 것으로 사료된다.

• 한국물환경학회지
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• 제20권5호
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• pp.506-511
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• 2004

Maintaining dissolved oxygen (DO) at sufficiently low concentration in the aeration tank at a wastewater treatment plant (WWTP) is essential for reduction of the costs of operation and maintenance. On the other hand, the low DO level may result in adverse effect on the integrity of the activated sludge, A typical and disastrous outcome frequently experienced is the outgrowth of filamentous microorganisms, which is called as filamentous bulking, In addition to the traditional methods such as sludge settleability and microscopic observation of the culture, molecular techniques including polymerase chain reaction (PCR) amplification followed by 16S rRNA sequencing were applied to identify filamentous bacteria present in bulking sludge under a condition of low DO concentration, Two morphologically distinct groups, presumably consisting of Sphaerofilus nafans, and Eikelboom Type 1701 or Type 1851, were identified through microscopic observation. They were further confirmed by subsequent 16S rRNA sequencing. Dominant filamentous bacteria identified by the molecular techniques were consisted of three major groups. Sequences of partial 16S rRNA cloned showed that the filamentous bulking organisms were closely related to Eikelboom Type 021N and Eikelboom Type 1701, and Sphaerotilus natans, respectively. Molecular methods were found to possess a strong potential of direct examination of the microbial community of an activated sludge system.

• 대한의생명과학회지
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• 제9권3호
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• pp.139-144
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• 2003

Since water borne infection causes acute diseases and results in spread of diseases by secondary infection, the prevention is very important. Therefore, it is necessary to have a method that is rapid and effective to monitor pathogenic bacteria in drinking water. In this study, we employed a systematic method, Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) analysis, to develop an effective monitoring system for possible bacterial contaminants in drinking water. For this purpose, PCR primers were derived from 992 bp region of the 16s rRNA gene that is highly conserved through the different species of prokaryotes. To test whether the PCR primers designed are indeed useful for detecting all the possible microbial contaminants in the water, the primers were used to amplify 16s rRNA regions of different microbial water-borne pathogens such as E. coli, Salmonella, Yersinia, Listeria, and Staphylococcus. As expected, all of tested microorganisms amplified expected size of PCR products indicating designed PCR primers for 16s rRNA indeed can be useful to amplify all different microbial water-borne pathogens in the water. Furthermore, to test whether these 16s rRNA based PCR primers can detect bacterial populations present in the water, water samples taken from diverse sources, such as river, tap, and sewage, were used for amplification. PCR products were for then subjected for cloning into a T-vector to generate a library containing 16s rRNA sequences from various bacteria. With cloned PCR products, RFLP analysis was done using PCR products digested with restriction enzyme such as Hae III to obtain species-specific RFLP profiles. After PCR-RFLP, the bacterial clones which showed the same RFLP profiles were regarded as the same ones, and the clones which showed distinctive RFLP profiles were subsequently subjected for sequence analysis for species identification. By this PCR-RFLP analysis, we were able to reveal diverse populations of bacteria living in water. In brief, in unsterilized natural river water, over 60 different species of bacteria were found. On the other hand, no PCR products were detected in drinking tap-water. The results from this study clearly indicate that the PCR-RFLP-sequence analysis can be a useful method for monitoring diverse, perhaps pathogenic bacteria contaminated in water in a rapid fashion.