• 한국치위생학회지
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• 제13권5호
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• pp.889-900
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• 2013

Objectives : Molecular biology techniques were employed to assess diversity of bacterial in children's dental caries. Methods : DNA of germs was extracted and the diversity of the 16S rRNA clones was analyzed by amplified rDNA restriction analysis and sequencing. The experimental samples were pit and fissure caries (PC), deep dentinal caries (DC), smooth surface caries (SC), and supragingival plaque (PQ) from 50 children of age less than 12 years old. The control group was healthy teeth supragingival plaque (HT). Thirty clones from each 16S rRNA clone library of 5 samples were randomly selected, thus a total of 150 clones were analyzed. Results : Amplified rDNA restriction analysis uncovered 18, 20, 11, 17, and 22 phylotypes from healthy teeth, pit and fissure caries, deep dentinal caries, smooth surface caries, and supragingival plaque, respectively. Sequencing analysis found the dominance of Actinomycs naeslundii and Fusobacterium nucleatum in the healthy teeth; Leptotrichia sp. in the pit and fissure caries; Actinomyces sp., Streptococcus mutans, and Rahnella aquatilis in the deep dentinal caries; Streptococcus mutans and Actinomyces sp. in the smooth surface caries; Enterobacter hormaechei and Streptococcus sanguinis in the supragingival plaque. Conclusions : Clonal analysis identified 6 phyla, 20 genera, and 51 species.

• Journal of Microbiology and Biotechnology
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• 제27권7호
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• pp.1288-1299
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• 2017

Composting is widely used to transform waste into valuable agricultural organic fertilizer. Anaerobic ammonium-oxidizing (anammox) bacteria play an important role in the global nitrogen cycle, but their role in composting remains poorly understood. In the present study, the community structure, diversity, and abundance of anammox bacteria were analyzed using cloning and sequencing methods by targeting the 16S rRNA gene and the hydrazine oxidase gene (hzo) in samples isolated from compost produced from cow manure and rice straw. A total of 25 operational taxonomic units were classified based on 16S rRNA gene clone libraries, and 14 operational taxonomic units were classified based on hzo gene clone libraries. The phylogenetic tree analysis of the 16S rRNA gene and deduced HZO protein sequences from the corresponding encoding genes indicated that the majority of the obtained clones were related to the known anammox bacteria Candidatus "Brocadia," Candidatus "Kuenenia," and Candidatus "Scalindua." The abundances of anammox bacteria were determined by quantitative PCR, and between $2.13{\times}10^5$ and $1.15{\times}10^6$ 16S rRNA gene copies per gram of compost were found. This study provides the first demonstration of the existence of anammox bacteria with limited diversity in cow manure composting.

• 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.

• 한국미생물·생명공학회지
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• 제23권4호
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• pp.384-389
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• 1995

Streptoalloteichus hindustanus ATCC 31219, a nebramycin complex producer, is similar to Streptomyeces tenebrarius in a viewpoint of resistance to a wide range of aminoglycoside antibiotics. S. tenebrarius has resistance mechanisms of 16s rRNA methylation and aminogycoside modification. However, it is not known whether resistance mechanisms of Stall. hindustanus are the same as in S. tenebrarius. Therefore, we have tried to isolate resistance determinants from Stall. hindustanus. Two different types of aminoglycoside resistance determinants were isolated from Stall. hindustanus and expressed in Streptomyces lividans TK24. The apramycin resistance gene (amr) and the tobramycin resistance gene (tmr) isolated from Stall. hindustanus showed broad resistance spectrum against a dozen of aminoglycoside antibiotics. The complete nucleotide sequences of apramycin resistance gene (amr) were determined. The deduced amino acid sequence of the amr gene of Stall hindustanus ATCC 31219 showed extensive sequence homology to the 16s rRNA methylase gene (kamB) of S. tenebrarius.

• Journal of Microbiology and Biotechnology
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• 제6권5호
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• pp.364-365
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• 1996

A region encoding the 16S rRNA was cloned by PCR from Streptomyces melanosporofaciens 7489 and sequenced by the chain-termination dideoxy sequencing method. A phylogenetic tree constructed by sequence alignment of 24 Streptomyces species suggests that there is little evolutionary distance between this strain and Streptomyces rimosus.

• Journal of Microbiology and Biotechnology
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• 제21권6호
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• pp.567-573
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• 2011

We investigated the phylogenetic diversity of ammoniaoxidizing bacteria (AOB) in Yellow Sea continental shelf sediment by the cloning and sequencing of PCR-amplified amoA and 16S rRNA genes. Phylogenetic analysis of the amoA-related clones revealed that the diversity of AOB was extremely low at the study site. The majority (92.7%) of amoA clones obtained belonged to a single cluster, environmental amoA cluster-3, the taxonomic position of which was previously unknown. Phylogenetic analysis on AOB-specific 16S rRNA gene sequences also demonstrated a very low diversity. All of the cloned 16S rRNA gene sequences comprised a single phylotype that belonged to the members of uncultured Nitrosospira cluster-1, suggesting that AOB belonging to the uncultured Nitrosospira cluster-1 could carry amoA sequences of environmental amoA cluster-3.

• 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.

• 미생물학회지
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• 제49권4호
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• pp.415-418
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• 2013

차세대 염기서열 분석(next generation sequencing, NGS) 기술의 발전으로 16S rRNA의 염기서열 분석이 미생물 군집 분석의 주된 방법으로 사용되고 있다. 인간의 건강과 질병에 관여하는 미생물들을 밝혀내기 위한 인간 미생물군집 프로젝트(human microbiome project, HMP)가 최근에 완료되었다. HMP는 세균에 의해 발생하는 여러 질병들의 특성들을 밝혀내었고, 특히 장에 서식하는 세균들에 대해 많은 연구가 수행되었다. 비록 인간의 장내 세균들에 대한 연구는 잘 수행되어왔지만, 다른 가축의 장내 세균에 대한 연구는 거의 이루어지지 않았다. 본 연구에서는 가축의 분변 미생물 다양성에 관해 조사하였고, 분변미생물 생태연구의 중요성을 제시 할 것이다. 한국에서의 분변 미생물 군집 프로젝트(fecal microbiome project) 시작을 본 연구논문을 통해 보고하고자 한다.

• KSBB Journal
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• 제19권1호
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• pp.72-77
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• 2004

건강에 대한 관심의 증대로 인한 저염식품의 개발 필요성에 맞추어 오이발효에 염의 농도를 낮추게 되면 정상적 일차발효 후에 이차발효가 진행됨으로써 결국 부패하게 된다. 장기간에 걸쳐 다양한 균의 복합적 작용으로 진행되는 저염 발효오이의 부패 과정을 이해하기 위하여 이에 관련된 균을 분리 동정하였다. 부패발효액을 무기배양하여 균을 분리하고 이들의 16s rRNA 유전자 부분을 universal primer를 이용한 PCR로서 증폭하여 서열분석 하였다. 분석된 800 염기 길이의 서열 전체를 그대로 이용하여 NCBI의 BLAST로서 유사종을 찾고 RDP의 Sequence Aligner와 Sequence Match에서 재확인하여 분리된 균을 3속 8종으로 동정하였다. Database 내의 표준균 서열을 기반으로 한 제한효소 지도와 동정된 균의 PCR 생성묵의 제한효소 처리결과(RFLP)를 비교하여 동정 결과를 실험으로 검정하였다. 동정과정에서 sequencing 결과 전체를 이용하는 점과 RDP를 통한 확인과 RFLP를 이용한 검정은 동정 결과에 대한 신뢰도를 한층 증가시켰다. 또 분리된 8종은 개별적 특징의 조사나 적절한 조합을 이룰 때의 상호 의존도 등을 조사할 수 있게 함으로써 여러 균에 의한 복합적 과정인 부패를 순차적 혹은 요인별로 나누어 살펴보는 연구를 가능하게 한다.

• 대한수의학회지
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• 제58권1호
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• pp.51-55
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• 2018

Bovine mastitis (BM) has resulted in enormous economic loss in the dairy industry and coagulase-negative staphylococci (CNS) have caused subclinical BM. Although VITEK 2 GP ID card (VITEK 2) has been used for CNS identification, the probability of identification varies. The rpoB sequence typing (RSTing) method has been used for molecular diagnosis and epidemiology of bacterial infections. In this study, we undertook RSTing of CNS and compared the results with those of VITEK2 and 16S rRNA gene sequencing. As compared VITEK2, the molecular-based methods were more reliable for species identification; moreover, RSTing provided more molecular epidemiological information than that from 16S rRNA gene sequencing.