• Bulletin of the Korean Chemical Society
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• 제30권11호
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• pp.2809-2811
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• 2009

• BMB Reports
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• 제48권12호
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• pp.643-644
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• 2015

In eukaryotes, small RNAs play important roles in both gene regulation and resistance to viral infection. Argonaute proteins have been identified as a key component of the effector complexes of various RNA-silencing pathways, but the mechanistic roles of Argonaute proteins in these pathways are not clearly understood. To address this question, we performed single-molecule fluorescence experiments using an RNA-induced silencing complex (core-RISC) composed of a small RNA and human Argonaute 2. We found that target binding of core-RISC starts at the seed region of the guide RNA. After target binding, four distinct reactions followed: target cleavage, transient binding, stable binding, and Argonaute unloading. Target cleavage required extensive sequence complementarity and accelerated core-RISC dissociation for recycling. In contrast, the stable binding of core-RISC to target RNAs required seed-match only, suggesting a potential explanation for the seed-match rule of microRNA (miRNA) target selection.

• 자연과학논문집
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• 제14권1호
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• pp.39-50
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• 2004

박테리오 파아지 T7 RNA 중합효소는 다른 RNA 중합효소와 비교하여 볼 때 보조인자 없이 전사를 진행하는 하나의 subunit로 구성된 RNA 중합효소이다. 전사 진행 단계 중에서 T7 RNA 중합효소의 전사연장을 연구하기 위해 biotin이 결합된 DNA 주형을 streptavidin bead로 고정시킴으로서 T7 RNA 중합효소의 진행과정을 관찰할 수 있었고, 이러한 기작을 이용하여 일련의 활성을 가지는 가장 안정한 전사연장복합체들을 얻을 수 있었다. 전사 연장체들은 16번 염기 위치로부터 18번 염기의 위치까지 방사선 동위원소가 표지되어 있으며 이들 표지된 전사연장복합체들은 단계별로 합성하여 22-40개 핵산잔기들이 합성된 전사연장복합체들을 얻을 수 있었다. 이와 같은 전사연장복합체들을 PTH 전사종결 부위가 있는 주형으로 사용하여 야생형 및 R173C 돌연변이 RNA 중합효소를 이용하여 전사연장복합체를 제조하여 비교한 결과 PTH 전사종결에 둔감한 R173C 돌연변이 중합효소의 경우 야생형에 비해 PTH 전사종결부위를 지난 위치에서도 전사연장복합체가 생성되었다.

• 대한미생물학회지
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• 제3권1호
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• pp.51-65
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• 1968

The site of the synthesis of Japanese encephalitis virus(JEV) in the actinomycin-treated and infecter PS Y15 cells(a porcine kidney stable cell line) was observed by the immunofluorescent antibody technique, acridine orange staining, and the autoradiographic analysis. In the parallel studies by immunofluorescent technique and acridine orange staining it the infected cells, Viral protein(as an antigen) and viral RNA were detected at the same site of cytoplasm. In the autoradiographic analysis, the cytoplasmic labeling of $^3H$-uridine was due to the synthesis of JEV-RNA, while the nucleolus and nucleus were not involved. In the autoradiographic studies on the secton of infected cells, the $^3H$-uridine was frequently incorporated around the cytoplasmic vacuoles. This localization of labeling agreed with the site of acridine orange positive granules. The results suggest that the syntheses of the viral RNA and viral protein occurred in the similar site of cytoplasm of the infected cells, and also the virus particles seem to be assembled in the sites of the viral RNA and protein syntheses.

• 한국동물학회지
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• 제20권1호
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• pp.9-18
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• 1977

RNA 분해효소에 저항하는 RNA 분자들이 양서류의 난에 존재하는지의 여부를 조사하기 전에 필요한 몇가지 예비실험을 하기 위하여 Xenopus laevis에 난에서 RNA를 추출 하였다. Sephadex G-100 column chromatography는 세 개의 peak을 항상 보여주고 있다. 첫째 peak에 포함되어있는 고분자량의 RNA만을 $^{3}H$-dimethyl sulfate를 사용하여 시험관내에서 label하여 tRNA로부터의 base paired oligonucleotide의 참여를 배제하였다. 이 방법으로 아주 높은 specific activity를 얻을 수 있었으며 또한 부착된 methyl group은 대단히 안정성을 보였다.

• The Plant Pathology Journal
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• 제17권3호
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• pp.115-122
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• 2001

Potato virus X (PVX) is a flexuous rod-shaped virus containing a single plus-strand RNA. Viral RNA synthesis is precisely regulated by regulatory viral sequences and by viral and/or host proteins. RNA sequence element as well as stable RNA stem-loop structure in the 5' end of the genome affect accumulation of genomic RNA and subgenomic RNA (sgRNA). The putative sgRNA promoter regions upstream of the PVX triple gene block (TB) and coat protein (CP) gene were critical for both TB and CP sgRNA accumulation. Mutations that disrupted complementarity between a region at the 5' end of the genomic RNA and the sequences located upstream of each sgRNA initiation site is important for PVX RNA accumulation. Compensatory mutations that restore complementarity restored sgRNA accumulation levels. However, the extent of reductions in RNA levels did not directly correlate with the degree of complementarity, suggesting that the sequences of these elements are also important. Gel-retardation assays showed that the 5' end of the positive-strand RNA formed an RNA-protein complex with cellular proteins, suggesting possible involvement of cellular proteins for PVX replication. Future studies on cellular protein binding to the PVX RNA and their role in virus replication will bring a fresh understanding of PVX RNA replication.

• BMB Reports
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• 제29권2호
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• pp.133-136
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• 1996

The affinity cleavage reagent Methidiumpropyl-EDTA-Iron(II) is applied to the structural analysis of 5S rRNA. Analysis of cleavage sites induced by MPE-Fe(II) on 5S rRNA shows that MPE intercalates easily between the unstable base pairs or into the bulges, thereby it strongly cuts the nucleosides nearby. The stable helical stems A, B, D and E as well as loop d are weakly cut. Most of the single-stranded loops are not cleaved. Based on the cleavage pattern of the 5S rRNA by MPE-Fe(II) and RNase V1, we suggest that MPE-Fe(II) may be used as a potential chemical probe in searching for the unstable helical regions of RNA, and for the sequences that appear to be involved in folding and distorting 5S rRNA.

• Asian Pacific Journal of Cancer Prevention
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• 제14권9호
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• pp.5391-5396
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• 2013

Aim: To investigate the role of Golgi phosphoprotein 3 (GOLPH3) in tumour growth and metastasis of esophageal squamous cancer. Methods: A lentiviral shRNA-vector was utilized to stably knockdown GOLPH3 in Eca-109 esophageal squamous cancer cells. mRNA transcription and protein expression of GOLPH3 were examined by real-time quantitative PCR and Western blotting, respectively. Cell proliferation activity was assessed by MTT assay and invasion and migration potentials by matrigel invasion and transwell motility assays. Results: Stable knockdown in the GOLPH3 cell line was established. PD-A gene expression was significantly suppressed by lentivirus-mediated RNAi, which resulted in reducing the capacity for cell proliferation, migration, invasion and adhesion in vitro. In vivo, GOLPH3 depletion resulted in inhibition of tumour growth, with stable decrease in the expression of GOLPH3 in tumor xenografts. Conclusions: Our findings suggest that lentivirus mediated silencing of the GOLPH3 gene has a significant anti-tumour effect on esophageal squamous cancer in vitro and in vivo. In addition, the results indicate that GOLPH3 might be an effective molecular target for gene therapy in esophageal squamous cancer.

• 한국자원식물학회지
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• 제20권6호
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• pp.511-517
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• 2007

A stable method for strain distinction using viral RNA 1 structures analyses was developed and compared with the combined RT-PCR and RAPD methods. Seven out of 61 random primers were found to be polymorphic based on RAPD analysis resulting on the differentiation of the 33 BaYMV isolates into four distinct groups according to geographical districts. The first and largest group includes 13 isolate and consists mainly of two-rowed malting barley in Haenam area. The second group had ten collections from inland in west southern. The third group had seven isolates from west southern coastal region, where mainly six-rowed naked barley is cultivated. The last fourth group included three isolates from Gyungnam region in east southern area. Conclusively, RNA 1 analysis proved to be stable and efficient method for strain distinction for Korean BaYMV isolates. Further, results of pathogenicity and RNA 1 structure analyses revealed four groups BaYMV strains and were distributed all over Korea, represented by Naju, Haenam-okcheon, Iksan and Milyang.

• Journal of Microbiology and Biotechnology
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• 제27권1호
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• pp.112-121
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• 2017

Edwardsiella tarda is a gram-negative pathogenic bacterium in aquaculture that can cause hemorrhagic septicemia in fish. Many secreted proteins have already been identified as virulent factors of E. tarda. Moreover, since virulent phenotypes are based on the expression regulation of virulent genes, understanding the expression profile of virulent genes is important. A quantitative RT-PCR is one of the preferred methods for determining different gene expressions. However, this requires the selection of a stable reference gene in E. tarda, which has not yet been systematically studied. Accordingly, this study evaluated nine candidate reference genes (recA, uup, rpoB, rho, topA, gyrA, groEL, rpoD, and 16S rRNA) using the Excel-based programs BestKeeper, GeNorm, and NormFinder under different culture conditions. The results showed that 16S rRNA was more stable than the other genes at different culture growth phases. However, at the same culture time, topA was identified as the reference gene under the conditions of different strains, different culture media, and infection, whereas gyrA was identified under the condition of different temperatures. Thus, in experiments, the expression of gapA and fbaA in E. tarda was analyzed by RT-qPCR using 16S rRNA, recA, and uup as the reference genes. The results showed that 16S rRNA was the most suitable reference gene in this analysis, and that using unsuitable reference genes resulted in inaccurate results.