• 미생물학회지
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• 제41권2호
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• pp.117-124
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• 2005

Rotifer와 병든 넙치 자어로부터 분리된 2개의 균주는 표현형적인 특성 확인 결과 Vibrio ichthyoenteri로 확인이 되었다. V. ichthyoenteri를 검출하기 위래 고감도 PCR 방법 개발을 하기 위래 V. ichthyoenteri 16S-23S rRNA intergenic spacer region(ISR)을 분석하였고, V. ichthyoenteri중 특이적 primer를 개발하였다. V. ichthyoenteri 의 ISR를 분석한 결과 1개의 다형성 ISR type서열을 포함하고 있었다. ISR서열은 길이는 348bp이며 tRNA gene을 가지고 있지 않았다. 이 서열을 가지고 이미 알려진 다른 Vibrio 종의 ISR 서열과 mutiple alignment를 수행한 결과 여러 영역에서 높은 가변성을 나타내어 가변 부위를 표적으로 하여 V. ichthyoenteri를 검출하기 위한 종 특이적 primer를 제작하였다. 제작된 primer의 특이성을 확인하기 위해 Vibrio 표준균주 19종의 genomic DNA와 분리균주 18 group에 genomic DNA 그리고 V. ichthyoenteri와 가장 유사한 서열을 가지고 있다고 알려진 Vibrio 종의 genomic DNA를 가지고 시험하였다. 그 결과 본 연구에서 제작된 종 특이적 primer를가지고 PCR 반응을 하면 V. ichthyoenteri를 검출 할 수가 있다.

• 한국동물위생학회지
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• 제37권3호
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• pp.157-164
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• 2014

Clostridium chauvoei is the etiologic agent of blackleg, a high mortality rated disease infection mainly cattle. In the present study, the partial sequences of 16S rRNA and flagellin gene of C. chauvoei isolated in Jeonbuk, Korea were determined and compared with those of reference strain. Oligonucleotide primers were designed to amplify a 811 bp fragment of 16S rRNA gene and 1229 bp fragment of flagellin gene. Sequencing analysis of 16S rRNA gene showed high homology to the reference strains ranging 82.3% to 100%, while flagellin gene were different from published foreign clostridia, showing 98.7% to 72.0% nucleotide sequence homology. Phylogenetic analysis based on 16S rRNA gene revealed the close phylogenetic relationship of C. chauvoei and C. septicum in cluster I, which includes C. carnis, C. tertium, C. quinii, C. celatum, C. perfringens, C. absonum, C. botulinum B. Phylogentic analysis also revealed that flagellin gene formed a single cluster with C. chauvoei, C. septicum, C. novyi A, C. novyi B, C. tyrobutylicum, C. acetobutylicum. The genetic informations obtained from this study could be useful for the molecular study of C. chauvoei.

• 미생물학회지
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• 제43권3호
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• pp.232-235
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• 2007

해양 및 토양에서의 암모니아 산화는 세균에 비해 Crenarchaeota 그룹의 archaea에 의해 우세하게 일어나고 있음이 알려졌다. 서해 갯벌에서, 배양에 의존하지 알고, 특정 암모니아 산화유전자(amoA)를 가진 archaea을 동정하고자 디지털 PCR법을 응용한 nested PCR법을 개발하였다. amoA와 16S rRNA유전자가 동시에 증폭된 샘플의 분석결과, 16S rRNA유전자에 비해 amoA 유전자의 다양성 이 높았으며, I.1a 그룹의 crenarchaea가 I.1b 그룹의 crenarchaea보다 갯벌지역에서 암모니아 산화에 우점적으로 기여하고 있음을 알 수 있었다. 본 연구에서 시도된, 디지털 PCR과 multiplex-nested PCR을 접목한 접근법을 이용하면 특정 기능유전자를 가진 미생물을 환경에서 검증하는데 응용할 수 있을 것이다.

• 한국축산시설환경학회지
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• 제21권3호
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• pp.93-98
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• 2015

The objective of this study was to review application of next-generation sequencing (NGS) to investigate microbiome in the livestock sector. Since the 16S rRNA gene is used as a phylogenetic marker, unculturable members of microbiome in nature or managed environments have been investigated using the NGS technique based on 16S rRNA genes. However, few NGS studies have been conducted to investigate microbiome in the livestock sector. The 16S rRNA gene sequences obtained from NGS are classified to microbial taxa against the 16S rRNA gene reference database such as RDP, Greengenes and Silva databases. The sequences also are clustered into species-level OTUs at 97% sequence similarity. Microbiome similarity among treatment groups is visualized using principal coordinates analysis, while microbiome shared among treatment groups is visualized using a venn diagram. The use of the NGS technique will contribute to elucidating roles of microbiome in the livestock sector.

• 미생물학회지
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• 제55권1호
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• pp.1-8
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• 2019

Due to the major role played by several species of Streptococcus in the etiology of periodontitis, it is important to assess the pattern of Streptococcus pathogenic pathways within the infected subgingival pockets using a bacterial specific 16S rRNA fragment. From the total of 50 patients with periodontitis included in the study, only 23 Streptococcal isolates were considered for further analyses, in which their 16S rRNA fragments were amplified and sequenced. Then, a comprehensive phylogenetic tree was constructed and in silico prediction was performed for the observed Streptococcal species. The phylogenetic analysis of the subgingival Streptococcal species revealed a high discrimination power of the 16S rRNA fragment to accurately identify three groups of Streptococcus on the species level, including S. salivarius (14 isolates), S. anginosus (5 isolates), and S. gordonii (4 isolates). The employment of state-of-art in silico tools indicated that each Streptococcal species group was characterized with particular transcription factors that bound exclusively with a different 16S rRNA-based secondary structure. In conclusion, the observed data of the present study provided in-depth insights into the mechanism of each Streptococcal species in its pathogenesis, which differ in each observed group, according to the differences in the 16S rRNA secondary structure it takes, and the consequent binding with its corresponding transcription factors. This study paves the way for further interventions of the in silico prediction, with the main conventional in vitro microbiota identification to present an interesting insight in terms of the gene expression pattern and the signaling pathway that each pathogenic species follows in the infected subgingival site.

• Journal of Microbiology and Biotechnology
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• 제18권2호
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• pp.194-198
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• 2008

A newly designed Cytophaga-Flavobacteria-specific 16S rRNA gene primer pair was employed to investigate the CF community structure in the Bering Sea, revealing a previously unknown and unexpected high CF diversity in this high latitude cold sea. In total, 56 clones were sequenced and 50 unique CF 16 rRNA gene fragments were obtained, clustering into 16 CF subgroups, including nine cosmopolitan subgroups, five psychrophilic subgroups, and two putatively autochthonous subgroups. The majority of sequences (82%) were closely related to uncultured CF species and could not be classified into known CF genera, indicating the presence of a large number of so-far uncultivated CF species in the Bering Sea.

• Journal of Yeungnam Medical Science
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• 제19권1호
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• pp.1-10
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• 2002

With the establishment of rapid sequence analysis of 16S rRNA and the recognition of its potential to determine the phylogenetic position of any prokaryotic organism, the role of 16S rRNA similarities in the present species definition in bacteriology need to be clarified. Comparative studies clearly reveal the limitations of the sequence analysis of this conserved gene and gene product in the determination of relationship at the pathogenic strain level for which DNA-DNA reassociation experiments still constitute the superior method. Since today the primary structure of 16S rRNA is easier to determine than hybridization between DNA strands, the strength of the sequence analysis is to recognize the level at which DNA pairing studies need to be performed, which certainly applies to similarities of 97% and higher.

• 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 and Biotechnology
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• 제17권7호
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• pp.1204-1207
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• 2007

Structural analyses have shown that nucleotides at the positions 770 and 771 of Escherichia coli 16S rRNA are implicated in forming one of highly conserved intersubunit bridges of the ribosome, B2c. To examine a functional role of these residues, base substitutions were introduced at these positions and mutant ribosomes were analyzed for their protein synthesis ability using a specialized ribosome system. The results showed requirement of a pyrimidine at the position 770 for ribosome function regardless of the nucleotide identity at the position 771. Sucrose gradient profiles of ribosomes revealed that the loss of protein-synthesis ability of mutant ribosome bearing a base substitution from C to G at the position 770 stems from its inability to form 70S ribosomes. These findings indicate involvement of nucleotide at the position 770, not 771, in ribosomal subunit association and provide a useful rRNA mutation that can be used as a target to investigate the physical interaction between 16S and 23S rRNA.

• 한국식품과학회지
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• 제30권3호
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• pp.655-659
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• 1998

우리나라 각지에서 젓갈류를 수집하여 혈전용해능을 나타내는 균주들을 분리하였다. 분리된 균주들의 활성을 측정한 결과 2.04 plasmin unit의 가장 우수한 혈전용해활성을 나타내는 균주를 선별하여 동정하였다. 균주의 형태학적, 배양학적 및 생화학적 성질을 조사한 결과 Bacillus pumilus로 나타났으나, 균체의 지방산조성을 분석하여 동정한 결과 Bacillus atropheus로 나타났다. 최종적 결론을 얻기 위해, 16S rRNA partial sequence(V3 region)를 결정하여 분석한 결과 Bacillus subtilis의 16S rRNA partial sequence와 동일한 것으로 나타나, Bacillus subtilis KJ-48로 명명하였다.